In vitro biosynthesis of plasma proteins under ischemic conditions of closed-circuit perfusion of healthy and intoxicated rabbit liver

We are elaborating on the kinetics and mechanisms of septic rabbit liver to de novo biosynthesize acute-phase response (APR) proteins under in vitro conditions of deepening ischemia in reference to their in vivo prevalence in serum and cerebrospinal fluids (CSF) collected at predetermined times. The significance of the data is interpreted as relevant to grafting cadaveric liver into end-stage liver diseased patients and APR-induced ischemic heart diseases (IHD). Hepatic APR was induced by CCl(4)-intubation, and the administration of cholera toxin (CT) or scorpion venom (SV), or both, to rabbits. Hepatic functional efficiency, in terms of biosynthesis of APR proteins in closed circuit perfusion of the isolated intoxicated liver with oxygenated saline or L-15 media paralleled the two-dimensional immunoelectrophoresis (2D-IEP) spectrum of APR serum proteins at time of liver isolation. We are suggesting: (a) in vitro biosynthesis of plasma proteins by isolated perfused liver is the result of in vivo decoded and retained APR inflammatory signals; and (b) decoded inflammatory signals are expressed not withstanding the perfusate's organic composition. Furthermore, 90 min of ischemic perfusion in saline or L-15 medium precipitated mitochondrial aberrations which resulted in further deterioration of de novo biosynthesis of APR plasma proteins. Regardless of the nature of the inflammatory stimuli, mitochondrial aberrations rendered the perfused organ a biologically inert tissue mass that was incapable of resuming biological function upon perfusion with oxygenated L-15 medium. This is most likely due to ischemia-induced irreversible hepatic necrosis. Thus, in vitro aberrations of mitochondrial function(s) critically limit the capability of the isolated liver to resume its organic function to sustain biosynthesis of de novo plasma proteins. Extrapolation of these results to the surgical management of end-stage liver diseases points to the importance of the status and the handling protocol(s) of the cadaver donor liver prior to successful grafting. We conclude that although histology of a cadaver liver may reveal well-preserved hepatic cellular organelles with at least minimal intra- and intercellular communication required for viable hepatic function, we deem it essential to further define acceptable minimal capabilities to de novo biosynthesize plasma proteins by a cadaver liver as a measure of its functional viability and suitability for transplantation. Ultimately, this measure may improve the success of liver transplants with minimal surgical and drug interventions.     

妊娠期血清微量元素的变化

目的探讨妊娠期血清微量元素的变化规律。方法以原子吸收分光光度法分别测定421例孕妇和128例正常对照妇女血清铜(Cu)、锌(Zn)、铁(Fe)等微量元素浓度和血清镁(Mg)、钙(Ca)浓度。结果血清铜(Cu)在整个孕期内无显著性变化,P>0.05;血清锌(Zn)、铁(Fe)、镁(Mg)、钙(Ca)随孕期的增加而降低,P均<0.05。结论孕妇在妊娠期内应根据不同孕期,合理补充微量元素。     

磁场对实验性AMI大鼠血浆cAMP、cGMP和血清Mg~(2 )含量变化的影响

本实验将大鼠随机分为五组：空白对照组、磁作用对照组、ＡＭＩ组、ＡＭＩ药物（心得安）治疗组、ＡＭＩ磁场治疗组。分别采用放射免疫方法和原子分光光度法测定各组大鼠血浆ｃＡＭＰ、ｃＧＭＰ和血清Ｍｇ２＋含量。结果表明：ＡＭＩ磁场治疗组血浆ｃＡＭＰ、ｃＧＭＰ含量及ｃＡＭＰ／ｃＧＡＰ比值均明显低于ＡＭＩ组（Ｐ＜０．０５）,与药物治疗组近似,略高于空白对照组、磁作用对照组,空白对照组略高于磁使用对照组,磁作用对照组最低,两组间无显著差异（Ｐ＞０．０５）。ＡＭＩ磁场治疗组血清Ｍｇ２＋含量明显高于ＡＭＩ组（Ｐ＜０．０５）,与药物治疗组近似,略低于空白对照组、磁作用对照组,空白对照组和磁作用对照组最高,两组间无显著差异（Ｐ＞０．０５）。提示：磁场对动物实验性ＡＭＩ心肌具有一定的保护作用。     

氟涂层镁铝合金的体外生物相容性研究简

目的 研究氟涂层镁铝合金的体外生物相容性。方法实验分为空白对照组（N组）、镁铝合金组（M组）、氟涂层镁铝合金组（F组）和阳性对照组（P组）4组。细胞毒性实验：将L929细胞在各组的DMEM浸提液中培养,光学显微镜下观察细胞生长状况。应用WST-1法测量光密度（OD）值。溶血实验：按GB-T16175-2008《医用有机硅材料生物学评价实验方法》第13部分《溶血试验》进行实验。测量各样本的OD值,计算溶血率。豚鼠最大剂量致敏实验,按照GBJ16886.10-2005（（医疗器械生物学评价》第10部分《刺激与迟发型超敏反应试验》进行实验。观察激发阶段去除贴附片后24、48、72h豚鼠皮肤致敏情况。结果各观察期F组的形态分级为0级,M组为4级。各组各观察期内OD值差异均有统计学意义（F=312.96,P=0.000）。第3天,实验组OD值均高于P组（1.050±0.065 vs 0.292±0.010）（P〈0.05）。第5天、第7天,F组与N组OD值（1.429±0.096 vs 1.622±0.156,0.928±0.040 vs 50.995±0.070）处于同一水平（P〉0.05）,均高于P组（0.270±0.015,0.281±0.006）（P〈0.05）。M组溶血率为68.3％,F组为0.8％。24h、48h和72h后N组、M组、F组皮肤均无红斑。结论氟涂层镁铝合金体外实验显示具有良好的生物相容性。     

Microstructure,mechanical and corrosion properties of Mg–Dy–Gd–Zr alloys for medical applications

In previous investigations, a Mg–10Dy (wt.%) alloy with a good combination of corrosion resistance and cytocompatibility showed great potential for use as a biodegradable implant material. However, the mechanical properties of Mg–10Dy alloy are not satisfactory. In order to allow the tailoring of mechanical properties required for various medical applications, four Mg–10(Dy + Gd)–0.2Zr (wt.%) alloys were investigated with respect to microstructure, mechanical and corrosion properties. With the increase in Gd content, the number of second-phase particles increased in the as-cast alloys, and the age-hardening response increased at 200 °C. The yield strength increased, while the ductility reduced, especially for peak-aged alloys with the addition of Gd. Additionally, with increasing Gd content, the corrosion rate increased in the as-cast condition owing to the galvanic effect, but all the alloys had a similar corrosion rate (～0.5 mm year^?1) in solution-treated and aged condition.     

Inhibition of ω-conotoxin-sensitive Ca2+ channel currents by internal Mg2+ in cultured rat cerebellar granule neurones

The effects of changing the intracellular concentrations of either free Mg²⁺ ions ([Mg²⁺]_i) or Mg²⁺-bound adenosine triphosphate ([Mg · ATP]_i) on Ca²⁺ channel currents were assessed in cultured rat cerebellar granule neurones using the whole-cell patch-clamp technique. Raising [Mg²⁺]_i from 0.06 mM to 1.0 mM inhibited Ca²⁺ channel currents by approximately 50%. The action of -conotoxin GVIA (-CgTX), a selective inhibitor of N-type Ca²⁺ channels was also investigated. With increasing [Mg²⁺]_i, the proportion of current irreversibly blocked by -CgTX was reduced, and was negligible (approximately 5 pA of current) in the presence of [Mg²⁺]_i values of 0.5 mM or greater. Block of the -CgTX-sensitive current accounted for the reduction in total current by concentrations of [Mg²⁺]_i to 0.5 mM. Raising [Mg²⁺]_i had no effect on the rate of decay of Ca²⁺ currents, but did produce a negative shift in current activation, possibly due to a non-specific interaction with negative surface charge. Altering [Mg · ATP]_i from 0.3 to 5.0 mM caused a twofold increase in the size of currents without affecting the proportion of current sensitive to -CgTX. [Mg²⁺]_i was also effective in inhibiting the Ca²⁺ channel current following potentiation by increasing [Mg · ATP]_i. These data suggest that -CgTX-sensitive current in these cells is selectively inhibited by internal Mg²⁺ whereas both -CgTX-sensitive and -resistant components of current are potentiated by internal Mg · ATP. The mechanism by which Mg²⁺ inhibits N-type channels is unclear, but may involve an open channel block.     

Mechanical and Biological Properties of a Biodegradable Mg‐Zn‐Ca Porous Alloy

Objectives

As promising alternative to current metallic biomaterials, the porous Mg scaffold with a 3‐D open‐pore framework has drawn much attention in recent years due to its suitable biodegradation, biocompatibility, and mechanical properties for human bones. This experiment's aim is to study the mechanical properties, biosafety, and osteogenesis of porous Mg‐Zn alloy.

Methods

A porous Mg‐2Zn‐0.3Ca (wt%) alloy was successfully prepared by infiltration casting, and the size of NaCl particles was detected by a laser particle size analyzer. The microstructure of the Mg‐2Zn‐0.3Ca alloy was characterized by the stereoscopic microscope and Sirion Field emission scanning electron microscope. X‐ray computerized tomography scanning (x‐CT) was used to create the 3‐D image. The degradation rate was measured using the mass loss method and the pH values were determined together. The engineering stress–strain curve, compressive modulus, and yield strength were tested next. The bone marrow stromal cells (BMSC) were cultured in vitro. The CCK‐8 method was used to detect the proliferation of the BMSC. Alkaline phosphatase (ALP) and alizarin red staining were used to reflect the differentiation effects. After co‐culturing, cell growth on the material's surface was observed by scanning electron microscope (SEM). The cell adhesion was tested by confocal microscopy.

Results

The obtained results showed that by using near‐spherical NaCl filling particles, the porous Mg alloy formed complete open‐cell foam with a very uniform size of pores in the range of 500–600 μm. Benefitting from the small size and uniform distribution of pores, the present porous alloy exhibited a very high porosity, up to 80%, and compressive yield strength up to 6.5 MPa. The degradation test showed that both the pH and the mass loss rate had similar change tendency, with a rapid rise in the early stage for 1–2 day's immersion and subsequently remaining smooth after 3 days. In vitro cytocompatibility trials demonstrated that in comparison with Ti, the porous alloy accelerated proliferation in 1, 3, 5, and 7 days (P < 0.001), and the osteogenic differentiation test showed that the ALP activity in the experimental group was significantly higher (P = 0.017) and has more osteogenesis nodules. Cell adhesion testing showed good osteoconductivity by more BMSC adhesion around the holes. The confocal microscopy results showed that cells in porous Mg‐based alloy had better cytoskeletal morphology and were larger in number than in titanium.

Conclusions

These results indicated that this porous Mg‐based alloy fabricated by infiltration casting shows great mechanical properties and biocompatibilities, and it has potential as an ideal bone tissue engineering scaffold material for bone regeneration.

     

Novel regimen through combination of memantine and tea polyphenol for neuroprotection against brain excitotoxicity

NMDA receptors are abundant, ubiquitously distributed throughout the brain, fundamental to excitatory neurotransmission, and critical for normal CNS function. However, excessive glutamate overstimulates NMDA receptors, leading to increased intracellular calcium and excitotoxicity. Mitochondrial dysfunction associated with loss of Ca(2+)homeostasis and enhanced cellular oxidative stress has long been recognized to play a major role in cell damage associated with excitotoxicity. In this experiment, we attempted to explore whether treatment with memantine (an NMDA receptor antagonist) and tea polyphenol (an antioxidant and anti-inflammatory agent), either alone or in combination, is effective in neuroprotection in a mouse excitotoxic injury model. Memantine (10 mg/kg/day), tea polyphenol (60 mg/kg/day), or a combination (memantine 5 mg/kg/day plus tea polyphenol 30 mg/kg/day) was administered by oral gavage for 2 consecutive days before causing excitotoxic injury. Mice received a 0.3-microL NMDA [335 mM (pH 7.2)] injection into the left striatum. Locomotor activity was assessed 24 hr before and after excitotoxic injury. Brain synaptosomes were harvested 24 hr after excitotoxic injury for assessment of Na(+), K(+)-ATPase and Mg(2+)-ATPase activity, reactive oxygen species production, mitochondrial membrane potential (Delta Psi m), mitochondrial reductase activity (MTT test), and Ca(2+)concentration. The results showed that treatment with memantine could significantly rescue mitochondrial function by attenuating the decreased mitochondrial membrane potential (Delta Psi m) and mitochondrial reductase activity in mouse excitotoxic injury. Treatment with tea polyphenol could significantly decrease the increased production of synaptosomal reactive oxygen species (ROS) and thus reduced the deteriorative ROS-sensitive Na(+), K(+)-ATPase and Mg(2+)-ATPase activity. However, neither memantine nor tea polyphenol alone could significantly improve the impaired locomotor activity unless treatment was combined. Combined treatment with memantine and tea polyphenol could significantly protect mice against excitotoxic injury by reducing the increased synaptosomal ROS production, attenuating the decreased Na(+), K(+)-ATPase and Mg(2+)-ATPase activity, the mitochondrial membrane potential (Delta Psi m), the mitochondrial reductase activity, and the increased synaptosomal Ca(2+)concentration. In addition, the impairment in locomotor activity was also significantly improved. Therefore, the combined treatment of memantine and tea polyphenol is more effective in neuroprotection than either memantine or tea polyphenol alone in mouse excitotoxic injury. These findings provide useful information about the potential application of memantine and tea polyphenols in preventing clinical excitotoxic injury such as brain trauma, brain ischemia, epilepsy, and Alzheimer's disease.     

Do Interictal Discharges Promote or Control Seizures? Experimental Evidence from an In Vitro Model of Epileptiform Discharge

Summary: Interictal and ictal discharges are recorded from limbic structures in temporal lobe epilepsy patients. In clinical practice, interictal spikes are used to localize the epileptogenic area, but they also are assumed to promote ictal events. Here I review data obtained from combined slices of mouse hippocampus–entorhinal cortex that indicate an inverse relation between interictal and ictal events. In this preparation, application of 4-aminopyridine or Mg²⁺-free medium induce (a) interictal discharges that originated from CA3 and propagate (via the Schaffer collaterals) to CA1 and entorhinal cortex, to return to the hippocampus through the dentate area; and (b) ictal discharges that initiate in the entorhinal cortex and propagate to the hippocampus via the dentate gyrus. Interictal activity occurs throughout the experiment (up to 6 h), whereas ictal discharges disappear after 1–2 h. Schaffer collateral cut abolishes interictal discharges in CA1, entorhinal cortex, and dentate and reestablishes entorhinal ictal discharges. Moreover, ictal discharge generation in the entorhinal cortex after Schaffer collateral cut is prevented by mimicking CA3 activity with rhythmic electrical stimulation of CA1 outputs. Thus hippocampal interictal activity controls the ability of the entorhinal cortex to generate seizures. It also may be proposed that Schaffer collateral cut may model the epileptic condition in which CA3 damage results in loss of hippocampal control over the entorhinal cortex. In conclusion, these experiments demonstrate that interictal activity controls rather than promotes ictal events, and functional integrity of CA3 constitutes a critical control mechanism in temporal lobe epilepsy.