Fungal Enzyme l-Lysine α-Oxidase Affects the Amino Acid Metabolism in the Brain and Decreases the Polyamine Level

The fungal glycoprotein l-lysine α-oxidase (LO) catalyzes the oxidative deamination of l-lysine (l-lys). LO may be internalized in the intestine and shows antitumor, antibacterial, and antiviral effects in vivo. The main mechanisms of its effects have been shown to be depletion of the essential amino acid l-lys and action of reactive oxidative species produced by the reaction. Here, we report that LO penetrates into the brain and is retained there for up to 48 h after intravenous injection, which might be explained by specific pharmacokinetics. LO actively intervenes in amino acid metabolism in the brain. The most significant impact of LO was towards amino acids, which are directly exposed to its action (l-lys, l-orn, l-arg). In addition, the enzyme significantly affected the redistribution of amino acids directly associated with the tricarboxylic acid (TCA) cycle (l-asp and l-glu). We discovered that the depletion of l-orn, the precursor of polyamines (PA), led to a significant and long-term decrease in the concentration of polyamines, which are responsible for regulation of many processes including cell proliferation. Thus, LO may be used to reduce levels of l-lys and PA in the brain

Моделирование интернализации водорастворимых противоопухолевых цитостатиков в тонкой кишке экспресс-методом ex vivo с помощью хемилюминесценции

Introduction. The ability of the small intestine (internalization) to absorb water-soluble anticancer cytostatics determines the possibility of their oral administration. The ex-vivo express method that simulates the internalization of substances using a modified technique of an isolated «inverted» segment of the rat small intestine with flash chemiluminescence is adequate to solve the problem. Objectives: to evaluate the absorption of the new water-soluble anticancer cytostatics with different properties from the rat small intestine for preclinical study by oral administration. Material and Methods. Conjugated with acridinium (Acridinium NHS Ester, Toronto Research Chemicals, Canada) cytostatics were studied: low molecular weight (1) Anthrafuran-Acridinium (MW 0.8 kDa) and high molecular weight (2) Aimpila-Acridinium (MW 105 kDa) and (3) L-lysine-a-oxidase (LO-Acridinium, MW 122 kDa). Absorption was determined in a modified model of an isolated «inverted» segment of the rat small intestine using flash-chemiluminescence with the calculation of the relative light units (RLU). Results. It was shown that the absorption level of acridinium-conjugated cytostatics depending on molar concentration ranged from 55 % (1) to 1.7-11 % (2, 3) and 2500 (1) to 9.2-188 nmol/l (2, 3), respectively. The level of internalized Anthrafuran-Acridinium (55 %) was consistent with the known value of the effective non-conjugated cytostatic oral dose, which was two times higher than equitherapeutical parenteral dose: 100 mg/kg vs 50 mg/kg. Conclusion. The data obtained allow us to consider ex vivo express method for preclinical study of the various water-soluble anticancer cytostatics for screening and identification of an opportunity for oral administration and estimation of starting dose. The method has a good correlation with in vivo tests and economically favorable due to a quick response and small number of the tested agent.Введение. Способность к всасыванию в тонкой кишке (интернализация) водорастворимых противоопухолевых цитостатиков определяет возможность их перорального применения. Экспресс-метод ex vivo, моделирующий интернализацию веществ в рамках модифицированной методики изолированного «вывернутого» отрезка тонкой кишки крысы с импульсной хемилюминесценцией, адекватен для решения поставленной задачи. Цель исследования - оценка всасывания в организм из тонкой кишки новых водорастворимых противоопухолевых цитостатиков с различными свойствами для доклинического изучения при пероральном введении. Материал и методы. В исследование включены конъюгированные с Акридином (Acridinium NHS Ester, Toronto Research Chemicals, Canada) цитостатики: низкомолекулярный (1) Антрафуран-Акридин (MW 0,8 кДа) и высокомолекулярные (2) Аимпила-Акридин (MW 105 кДа) и (3) L-лизин-а-оксидаза (ЛО-Акридин, MW 122 кДа). Всасывание определено в модифицированной модели изолированного «вывернутого» отрезка тонкой кишки крысы методом импульсной флэш-хемилюминесценции и пересчитано в процентах. Результаты. Показано, что в зависимости от молярной концентрациии от 2500 (1) до 9,2-188 нмоль/л (2, 3) уровень всасывания конъюгирован-ных с Акридином цитостатиков находится в диапазоне от 55 % (1) до 1,7-11 % (2, 3) соответственно. Уровень всасывания конъюгированного Антрафурана (55 %) согласуется с величиной известной эффективной пероральной дозы неконъюгированного цитостатика, которая была в два раза больше, чем эквитерапевтическая парентеральная доза: 100 мг/кг против 50 мг/кг. Заключение. Полученные данные позволяют рассматривать экспресс-метод ex vivo для скрининга возможности доклинического изучения различных водорастворимых противоопухолевых цитостатиков при пероральном введении с прогнозированием стартовой дозы. Метод адекватен тестам in vivo и экономически целесообразен в силу быстрого ответа и малого количества тестируемого агента

Photoluminescent Scaffolds Based on Natural and Synthetic Biodegradable Polymers for Bioimaging and Tissue Engineering

Non-invasive visualization and monitoring of tissue-engineered structures in a living organism is a challenge. One possible solution to this problem is to use upconversion nanoparticles (UCNPs) as photoluminescent nanomarkers in scaffolds. We synthesized and studied scaffolds based on natural (collagen—COL and hyaluronic acid—HA) and synthetic (polylactic-co-glycolic acids—PLGA) polymers loaded with β-NaYF4:Yb3+, Er3+ nanocrystals (21 ± 6 nm). Histomorphological analysis of tissue response to subcutaneous implantation of the polymer scaffolds in BALB/c mice was performed. The inflammatory response of the surrounding tissues was found to be weak for scaffolds based on HA and PLGA and moderate for COL scaffolds. An epi-luminescent imaging system with 975 nm laser excitation was used for in vivo visualization and photoluminescent analysis of implanted scaffolds. We demonstrated that the UCNPs’ photoluminescent signal monotonously decreased in all the examined scaffolds, indicating their gradual biodegradation followed by the release of photoluminescent nanoparticles into the surrounding tissues. In general, the data obtained from the photoluminescent analysis correlated satisfactorily with the histomorphological analysis

Photoluminescent Scaffolds Based on Natural and Synthetic Biodegradable Polymers for Bioimaging and Tissue Engineering

Non-invasive visualization and monitoring of tissue-engineered structures in a living organism is a challenge. One possible solution to this problem is to use upconversion nanoparticles (UCNPs) as photoluminescent nanomarkers in scaffolds. We synthesized and studied scaffolds based on natural (collagen—COL and hyaluronic acid—HA) and synthetic (polylactic-co-glycolic acids—PLGA) polymers loaded with β-NaYF4:Yb3+, Er3+ nanocrystals (21 ± 6 nm). Histomorphological analysis of tissue response to subcutaneous implantation of the polymer scaffolds in BALB/c mice was performed. The inflammatory response of the surrounding tissues was found to be weak for scaffolds based on HA and PLGA and moderate for COL scaffolds. An epi-luminescent imaging system with 975 nm laser excitation was used for in vivo visualization and photoluminescent analysis of implanted scaffolds. We demonstrated that the UCNPs’ photoluminescent signal monotonously decreased in all the examined scaffolds, indicating their gradual biodegradation followed by the release of photoluminescent nanoparticles into the surrounding tissues. In general, the data obtained from the photoluminescent analysis correlated satisfactorily with the histomorphological analysis

Antitumor Effect of Bleomycin Nanoaerosol in Murine Carcinoma Model

Bleomycin, which is widely used as an antitumor agent, possesses serious adverse effects such as pulmonary toxicity. Local nanoaerosol deposition for lung cancer treatment is a promising alternative to drug delivery to lung lesions. The aim of this work is to test the hypothesis that bleomycin nanoaerosol can be effectively used to treat multiple lung metastases. To obtain bleomycin nanoaerosol, an aerosol generator based on electrospray of a solution of a nonvolatile substance with gas-phase neutralization of charged aerosol particles was used. Lung metastases in murine Lewis lung carcinoma and B16 melanoma animal models were counted. The effect of inhaled bleomycin nanoparticles on the number and volume of metastases, as well as pulmonary side effects, was investigated. Using a mouse exposure chamber, the dose-dependent effect of inhaled bleomycin on tumor volume was evaluated in comparison with intraperitoneal administration. Bleomycin nanoaerosol reduced the volume of metastases and produced a higher antitumor effect at much lower doses. It has been established that long-term exposure to nanoaerosol with a low dose of bleomycin is capable of suppressing cancer cell growth. The treatment was well tolerated. In the lungs, minor changes were found in the form of focal-diffuse infiltration of the lung parenchyma

Bioactivity Features of a Zn-1%Mg-0.1%Dy Alloy Strengthened by Equal-Channel Angular Pressing

The structure, phase composition, corrosion and mechanical properties, as well as aspects of biocompatibility in vitro and in vivo, of a Zn-1%Mg-0.1%Dy alloy after equal-channel angular pressing (ECAP) were studied. The structure refinement after ECAP leads to the formation of elongated α-Zn grains with a width of ~10 µm and of Mg- and Dy-containing phases. In addition, X-ray diffraction analysis demonstrated that ECAP resulted in the formation of the basal texture in the alloy. These changes in the microstructure and texture lead to an increase in ultimate tensile strength up to 262 ± 7 MPa and ductility up to 5.7 ± 0.2%. ECAP slows down the degradation process, apparently due to the formation of a more homogeneous microstructure. It was found that the alloy degradation rate in vivo after subcutaneous implantation in mice is significantly lower than in vitro ones. ECAP does not impair biocompatibility in vitro and in vivo of the Zn-1%Mg-0.1%Dy alloy. No signs of suppuration, allergic reactions, the formation of visible seals or skin ulcerations were observed after implantation of the alloy. This may indicate the absence of an acute reaction of the animal body to the Zn-1%Mg-0.1%Dy alloy in both states

Biocompatibility and Degradation of Fe-Mn-5Si Alloy after Equal-Channel Angular Pressing: In Vitro and In Vivo Study

An attempt to improve the functional characteristics of a degradable Fe-Mn-5Si shape memory alloy by means of structure refinement by equal-channel angular pressing (ECAP) was made. In the course of ECAP, an austenitic ultrafine-grained structure was obtained. In shear bands with a thickness of 301 ± 31 nm, twins 11 ± 1 nm in size were formed. Due to the resulting structure, the tensile strength was doubled up to 1419 MPa, and the yield strength was increased up to 1352 MPa, four times higher compared with the annealed state. Dynamic indentation tests revealed a decrease in Young’s modulus by more than 2.5 times after ECAP compared to values measured in the annealed state. The results of the study of hemolytic and cytotoxic activity in vitro, as well as the local and systemic reactivity of the body of laboratory animals after implantation of the test samples indicate the biocompatibility of the alloy after ECAP. Biocompatibility, high specific strength and low modulus of elasticity open prospects for Fe-Mn-5Si alloy after ECAP to be used for the production of degradable implants that can effectively provide the fastening function in osteoreconstruction

Upconversion Nanoparticles Intercalated in Large Polymer Micelles for Tumor Imaging and Chemo/Photothermal Therapy

Frontiers in theranostics are driving the demand for multifunctional nanoagents. Upconversion nanoparticle (UCNP)-based systems activated by near-infrared (NIR) light deeply penetrating biotissue are a powerful tool for the simultaneous diagnosis and therapy of cancer. The intercalation into large polymer micelles of poly(maleic anhydride-alt-1-octadecene) provided the creation of biocompatible UCNPs. The intrinsic properties of UCNPs (core@shell structure NaYF4:Yb3+/Tm3+@NaYF4) embedded in micelles delivered NIR-to-NIR visualization, photothermal therapy, and high drug capacity. Further surface modification of micelles with a thermosensitive polymer (poly-N-vinylcaprolactam) exhibiting a conformation transition provided gradual drug (doxorubicin) release. In addition, the decoration of UCNP micelles with Ag nanoparticles (Ag NPs) synthesized in situ by silver ion reduction enhanced the cytotoxicity of micelles at cell growth temperature. Cell viability assessment on Sk-Br-3, MDA-MB-231, and WI-26 cell lines confirmed this effect. The efficiency of the prepared UCNP complex was evaluated in vivo by Sk-Br-3 xenograft regression in mice for 25 days after peritumoral injection and photoactivation of the lesions with NIR light. The designed polymer micelles hold promise as a photoactivated theranostic agent with quattro-functionalities (NIR absorption, photothermal effect, Ag NP cytotoxicity, and Dox loading) that provides imaging along with chemo- and photothermal therapy enhanced with Ag NPs