Effect of taktivin on diuresis and tubular cardiotrast secretion in the kidney

The effect of a new immunomodulator taktivin (7-20 micrograms/kg subcutaneously, for 6 days) on the diuresis and tubular transport of cardiotrast (diodrast) was studied on rats. taktivin was shown to increase the tubular transport of the xenobiotic without significant changes in glomerular filtration and renal excretion of water, sodium, potassium, uric acid and creatinine. Possible mechanism of taktivin's action on the tubular transport of xenobiotics is discussed.     

The Hidden Cost of Housing Practices: Using Noninvasive Imaging to Quantify the Metabolic Demands of Chronic Cold Stress of Laboratory Mice

Laboratory mice routinely are housed at 20 to 22 °C—well below the murine thermoneutral zone of 29 to 34 °C. Chronic cold stress requires greater energy expenditure to maintain core body temperature and can lead to the failure of mouse models to emulate human physiology. We hypothesized that mice housed at ambient temperatures of 20 to 22 °C are chronically cold-stressed, have greater energy expenditure, and have high glucose utilization in brown adipose tissue. To test our hypotheses, we used indirect calorimetry to measure energy expenditure and substrate utilization in C57BL/6J and Crl:NU-Foxn1^nu nude mice at routine vivarium (21 °C), intermediate (26 °C), and heated (31 °C) housing temperatures. We also examined the activation of interscapular brown adipose tissue, the primary site of nonshivering thermogenesis, via thermography and glucose uptake in this region by using positron emission tomography. Energy expenditure of mice was significantly higher at routine vivarium temperatures compared with intermediate and heated temperatures and was associated with a shift in metabolism toward glucose utilization. Brown adipose tissue showed significant activation at routine vivarium and intermediate temperatures in both hirsuite and nude mice. Crl:NU-Foxn1^nu mice experienced greater cold stress than did C57BL/6J mice. Our data indicate mice housed under routine vivarium conditions are chronically cold stress. This novel use of thermography can measure cold stress in laboratory mice housed in vivaria, a key advantage over classic metabolic measurement tools. Therefore, thermography is an ideal tool to evaluate novel husbandry practices designed to alleviate murine cold stress.Abbreviations: BAT, brown adipose tissue; EPR, entropy production rate; PET, positron emission tomography; ROI, region of interest; RQ, respiratory quotient; VCO₂, volume of carbon dioxide produced; VO₂, volume of oxygen consumedLaboratory mice routinely are housed at ‘room temperature,’ that is, 20 to 22 °C. Room temperature is within the recommendations of the Guide for the Care and Use of Laboratory Animals¹⁴ but is well below the murine thermoneutral zone of 29 to 34 °C.^3,17 Systemic physiologic cold stress creates a much greater energy demand on mice than humans due to the surface area to volume ratio. Mice housed at routine vivarium temperatures have greater oxygen consumption and feed intake than at thermoneutral temperatures (30 °C).^5,27 Ultimately, this difference may adversely affect translational research, sometimes in unpredictable ways.^11,17 For example, mice housed at temperatures below their thermoneutral zone have a blunted response to LPS-induced fever and lack the classic early-phase hypothermia, demonstrating impaired immune function.²² In another example, blood pressure and heart rate are significantly elevated at routine vivarium temperatures compared with thermoneutral temperatures,²³ again demonstrating that rodent physiology is perturbed under such housing conditions.Mammals defend their body temperature through a series of mechanisms that progressively increase in energy cost: behavior,¹⁰ insulative response,⁷ and thermogenesis.³ Behavioral thermoregulation is the principle mechanism that enables the survival of small rodents, however behavioral adaptations of laboratory rodents housed in barren cages are limited compared with those of their wild counterparts; wild rodents adapt to cold stress through techniques like seeking shelter, burrowing, and building nests.^9,10 The insulative response shunts blood from peripheral sites toward core organs to conserve heat.⁷ Once the low-energy cold-adaptive responses are overwhelmed, mammals maintain core body temperature by increasing energy expenditure via shivering and nonshivering thermogenesis.^9,19 Rodents, arctic mammals, and infant mammals primarily rely on nonshivering thermogenesis to preserve core body temperature.²Nonshivering thermogenesis is achieved through mitochondria-rich brown adipose tissue (BAT). The largest deposits of rodent BAT are located in the interscapular region. BAT produces heat rapidly via the oxidative combustion of glucose and triglycerides.^2,6 BAT is rich in β₃-adrenergic receptors, and its activation is mediated primarily by the sympathetic nervous system.² BAT responds within minutes to the sensation of cold. When active, rodent BAT is highly metabolic and can receive as much as 40% of the cardiac outflow.⁷Given our group''s experience with preclinical metabolic imaging,⁸ we hypothesized that the cold stress imposed by routine husbandry temperatures (21 °C) induces a global shift toward glucose-dependent metabolism that is driven by nonshivering thermogenesis. Moreover, we hypothesized that athymic nude (Crl:NU-Foxn1^nu) mice housed at room temperatures experience significantly greater energy expenditure and glucose-dependent metabolism than do hirsute (C57BL/6J) mice.To test our hypotheses, we measured energy expenditure (entropy production rate, EPR, cal/min) and metabolic substrate utilization via indirect calorimetry⁶ and interscapular BAT heat production via infrared thermography and BAT glucose utilization by fluorodeoxyglucose positron emission tomography (PET) at various environmental temperatures, ranging from routine vivarium temperature (21 °C) to heat-supported temperatures (31 °C). Briefly, indirect calorimetry measures O₂ consumption (VO₂) and CO₂ production (VCO₂) to enable the calculation of the energy expenditure of the organism according to the stoichiometric formulas of biologic combustion:With these formulas, indirect calorimetry can also be used to calculate global glucose and lipid utilization using the respiratory quotient (RQ), a unitless ratio between VCO₂:VO₂.⁵ An RQ of 0.7 is indicative of the use of lipid as the primary substrate of biologic combustion, whereas an RQ of 1.0 is indicative of the primary use of glucose (for additional information, see reference 6).     

A one step/one pot synthesis of N,N-bis(phosphonomethyl)amino acids and their effects on adipogenic and osteogenic differentiation of human mesenchymal stem cells

The one pot reaction of amino acids with diethylphosphite and formaldehyde yielded N,N-bis(phosphonomethyl)amino acids. This synthetic route does not require harsh reagents to cleave the ester group. The molecular structures of the new compounds were determined by X-ray diffraction methods. By employing DFT calculations the hydrolysis of the intermediate phosphonic esters to the respective acids could be explained by the decreasing P–OEt bond strength for C_α-bisalkylated amino acids. Biological evaluation on the adipogenic and osteogenic differentiation of mesenchymal stem cells revealed no modification of the adipocyte differentiation, but inhibition of osteoblast formation at concentrations without detectable cytotoxicity.     

Ribonucleotide reductase inhibition by metal complexes of Triapine (3-aminopyridine-2-carboxaldehyde thiosemicarbazone): a combined experimental and theoretical study

Triapine (3-aminopyridine-2-carboxaldehyde thiosemicarbazone, 3-AP) is currently the most promising chemotherapeutic compound among the class of α-N-heterocyclic thiosemicarbazones. Here we report further insights into the mechanism(s) of anticancer drug activity and inhibition of mouse ribonucleotide reductase (RNR) by Triapine. In addition to the metal-free ligand, its iron(III), gallium(III), zinc(II) and copper(II) complexes were studied, aiming to correlate their cytotoxic activities with their effects on the diferric/tyrosyl radical center of the RNR enzyme in vitro. In this study we propose for the first time a potential specific binding pocket for Triapine on the surface of the mouse R2 RNR protein. In our mechanistic model, interaction with Triapine results in the labilization of the diferric center in the R2 protein. Subsequently the Triapine molecules act as iron chelators. In the absence of external reductants, and in presence of the mouse R2 RNR protein, catalytic amounts of the iron(III)–Triapine are reduced to the iron(II)–Triapine complex. In the presence of an external reductant (dithiothreitol), stoichiometric amounts of the potently reactive iron(II)–Triapine complex are formed. Formation of the iron(II)–Triapine complex, as the essential part of the reaction outcome, promotes further reactions with molecular oxygen, which give rise to reactive oxygen species (ROS) and thereby damage the RNR enzyme. Triapine affects the diferric center of the mouse R2 protein and, unlike hydroxyurea, is not a potent reductant, not likely to act directly on the tyrosyl radical.     

Biomolecular strategies of bone augmentation in spinal surgery

Autologous bone grafts and allografts are the most accepted procedures for achieving spinal fusion. Recently, breakthroughs in understanding bone biology have led to the development of novel approaches to address the clinical problem of bone regeneration in an unfavorable environment, while bypassing the drawbacks of traditional treatments, including limited availability, donor site morbidity, risk of disease transmission and reduced osteogenicity. These approaches have also been studied for their effectiveness in reaching successful spinal fusion. This review focuses on the cellular and molecular mechanisms explaining the rationale behind these methods, including bone marrow aspirate and mesenchymal stem cells, platelet-rich plasma, bone morphogenetic proteins and gene therapy, which have opened a promising perspective in the field of bone formation in spinal surgery.     

M-phase-specific protein kinase from mitotic sea urchin eggs: cyclic activation depends on protein synthesis and phosphorylation but does not require DNA or RNA synthesis

Histone H1 kinase (H1K) undergoes a transient activation at each early M phase of both meiotic and mitotic cell cycles. The mechanisms underlying the transient activation of this protein kinase were investigated in mitotic sea urchin eggs. Translocation of active H1K from particulate to soluble fraction does not seem to be responsible for this activation. H1K activation cannot be accounted for by the transient disappearance of a putative H1K inhibitor present in soluble fractions of homogenates. Aphidicolin, an inhibitor of DNA synthesis, and actinomycin D, an inhibitor of RNA synthesis, do not impede the transient appearance of H1K activity. H1K activation therefore does not require DNA or RNA synthesis. Fertilization triggers a rise in intracellular pH responsible for the increase of protein synthesis. H1K activation is highly dependent on the intracellular pH. Ammonia triggers an increase of intracellular pH and stimulates protein synthesis and H1K activation. Acetate lowers the intracellular pH, decreases protein synthesis, and blocks H1K activation. Protein synthesis is an absolute requirement for H1K activation as demonstrated by their identical sensitivities to emetine concentration and to time of emetine addition. About 60 min after fertilization, H1K activation and cleavage become independent of protein synthesis. The concentration of p34, a homolog of the yeast cdc2 gene product which has been recently shown to be a subunit of H1K, does not vary during the cell cycle and remains constant in emetine-treated cells. H1K activation thus requires the synthesis of either a p34 postranslational modifying enzyme or another subunit. Finally, phosphatase inhibitors and ATP slow down in the in vitro inactivation rate of H1K. These results suggest that a subunit or an activator of H1K is stored as an mRNA in the egg before mitosis and that full activation of H1K requires a phosphorylation.     

The phosphohydrolase component of the hepatic microsomal glucose-6-phosphatase system is a 36.5-kilodalton polypeptide

The phosphohydrolase component of the microsomal glucose-6-phosphatase system has been identified as a 36.5-kDa polypeptide by 32P-labeling of the phosphoryl-enzyme intermediate formed during steady-state hydrolysis. A 36.5-kDa polypeptide was labeled when disrupted rat hepatic microsomes were incubated with three different 32P-labeled substrates for the enzyme (glucose-6-P, mannose-6-P, and PPi) and the reaction terminated with trichloroacetic acid. Labeling of the phosphoryl-enzyme intermediate with [32P]glucose-6-P was blocked by several well-characterized competitive inhibitors of glucose-6-phosphatase activity (e.g. Al(F)-4 and Pi) and by thermal inactivation, and labeling was not seen following incubations with 32Pi and [U-14C]glucose-6-P. In agreement with steady-state dictates, the amount of [32P]phosphoryl intermediate was directly and quantitatively proportional to the steady-state glucose-6-phosphatase activity measured under a variety of conditions in both intact and disrupted hepatic microsomes. The labeled 36.5-kDa polypeptide was specifically immunostained by antiserum raised in sheep against the partially purified rat hepatic enzyme, and the antiserum quantitatively immunoprecipitated glucose-6-phosphatase activity from cholate-solubilized rat hepatic microsomes. [32P]Glucose-6-P also labeled a similar-sized polypeptide in hepatic microsomes from sheep, rabbit, guinea pig, and mouse and rat renal microsomes. The glucose-6-phosphatase enzyme appears to be a minor protein of the hepatic endoplasmic reticulum, comprising about 0.1% of the total microsomal membrane proteins. The centrifugation of sodium dodecyl sulfate-solubilized membrane proteins was found to be a crucial step in the resolution of radiolabeled microsomal proteins by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.