Measurement of quantity of iron in magnetically labeled cells: comparison among different UV/VIS spectrometric methods

Cell labeling with superparamagnetic iron oxides (SPIO) is becoming a routine procedure in cellular magnetic resonance imaging (MRI). Quantifying the intracellular iron in labeled cells is a prerequisite for determining the number of accumulated cells by quantitative MRI studies. To establish the most sensitive and reproducible method for measuring iron concentration in magnetically labeled cells, we investigated and compared four different methods using an ultraviolet-visible (UV/VIS) spectrophotometer. Background spectra were obtained for 5 and 10 M hydrochloric acids, a mixture of 100 mM citric acid plus ascorbic acid and bathophenanthroline sulphonate (BPS), and a mixture of 5 M hydrochloric acid plus 5% ferrocyanide. Spectra of the same solutions containing either 10 or 5 microg/mL iron oxides were also created to determine the peak absorbance wavelengths for the dissolved iron. In addition, different known iron concentrations were used to obtain calibration lines for each method. Based on the calibration factors, iron was measured in samples with a known amount of iron and in labeled cells. Methods based on the use of 10 M hydrochloric acid underestimated iron concentration in all experiments; for this method to give an accurate measurement, iron concentration in sample needs to be at least 3 microg/mL.     

Voxel model of a rabbit: assessment of absorbed doses in organs after CT examination performed by two different protocols

The objective of this work was to assess absorbed doses in organs and tissues of a rabbit, following computed tomography (CT) examinations, using a dedicated 3D voxel model. Absorbed doses in relevant organs were calculated using the MCNP5 Monte Carlo software. Calculations were perfomed for two standard CT protocols, using tube voltages of 110 kVp and 130 kVp. Absorbed doses were calculated in 11 organs and tissues, i.e., skin, bones, brain, muscles, heart, lungs, liver, spleen, kidney, testicles, and fat tissue. The doses ranged from 15.3 to 28.3 mGy, and from 40.2 to 74.3 mGy, in the two investigated protocols. The organs that received the highest dose were bones and kidneys. In contrast, brain and spleen were organs that received the smallest doses. Doses in organs which are stretched along the body did not change significantly with distance. On the other hand, doses in organs which are localized in the body showed maximums and minimums. Using the voxel model, it is possible to calculate the dose distribution in the rabbit’s body after CT scans, and study the potential biological effects of CT doses in certain organs. The voxel model presented in this work can be used to calculated doses in all radiation experiments in which rabbits are used as experimental animals.

     

Unusual allometry between in situ growth of freshwater phytoplankton under static and fluctuating light environments: possible implications for dominance

The effects of fluctuating light fields on the growth of phytoplanktonare not well understood and conclusions in the literature havebeen equivocal. Most studies have examined responses such asproductivity and chlorophyll a content (laboratory culture andfield tests) or growth rates (laboratory culture tests). Inthis study we examined the in situ growth rates of differenttypes of phytoplankton within two natural populations. Comparisonswere made between populations grown in a static environment(suspended in a fixed position in the water column) and an equivalentpopulation moving through the water column simulating the mixingof entrained phytoplankton. Growth under fluctuating light fieldsin this experiment only significantly (P < 0.05) increasedthe growth of the diatom Skeletonema and decreased the growthof Anabaena circinalis, Microcystis aeruginosa and Scenedesmussp. All other phytoplankton, including the genera Nitzschia,Fragilaria and Dactylococcopsis, did not have growth rates thatwere significantly different between static and fluctuatinglight treatments. A general pattern where diatoms grew best,followed by chlorophytes with the toxicogenic cyanophytes M.aeruginosa and A. circinalis growing least well, was distinguishedunder fluctuating irradiance. This seems consistent with thecommon occurrence of these groups of phytoplankton in the naturalenvironment. The cyanophytes Dactylococcopsis and Aphanothecedid not follow this pattern, with the former growing betterunder fluctuating light and the latter exhibiting an unusualgrowth pattern where growth was higher under lower light intensities.     

Anin VitroModel of Oligodendrocyte Destruction by Nitric Oxide and Its Relevance to Multiple Sclerosis

There is mounting evidence that nitric oxide (NO) is produced in the brains of patients with multiple sclerosis (MS) and in the experimental model of MS, experimental autoimmune encephalomyelitis, after the induction of Type II nitric oxide synthase (iNOS). Because NO can cause a variety of biological insults that compromise or even kill normal cells, we studied the effects of NO on oligodendrocytes since they are a target in MS tissue. In anin vitromodel, we have been able to demonstrate that NO causes damage to oligodendrocytes preferentially, sparing microglia almost completely and affecting some but not all astrocytic functions. This article describes the types of assays used to measure morphological changes, mitochondrial dysfunction, DNA strand breaks, and cell death brought on by NO or peroxynitrite (ONOO-) as well as a comprehensive review of the various techniques and sensitivities of NO and iNOS assays that would be applicable to similarin vitromodels.     

Development of the diarthrodial joints in the rat embryo

The development of appendicular synovial joints of both legs was studied with histological and histochemical techniques in 43 rat embryos aged 12 to 21 days. From this and previous studies, it appears that joints develop by a sequence of cellular events leading to a full expression of the phenotypic characteristics. The classically described stages: cell condensation, three layered mesenchyme, vascular invasion and joint clefting, were chronologically recorded in all joints. The observations relevant to the intra-articular structures, such as joint capsule, menisci and ligaments, were also presented. Previously unreported, cellular aspects were described during joint morphogenesis and their biological significance was discussed. Among these cellular aspects, of particular interest are: a. an early wave of cell necrosis occurring immediately after differentiation of the interzone. Disappearance of necrotic cells is thought to prevent chondrification of this tissue by clearing up the cells with chondroblastic potentialities; and b. a morphologically peculiar type of cells that differentiate alongside, and by the time of, clefting and seem to be related to this process. Thus, the joint clefting appears also to result from a cell-tissue related phenomenon, acting in conjunction with the joint motion, the importance of which has been previously demonstrated.     

C-kit-immunopositive interstitial cells of Cajal in human embryonal and fetal oesophagus

Interstitial cells of Cajal (ICC) are morphologically and functionally intercalated between the elements of the enteric nervous system and the smooth muscle cells (SMCs) in the musculature of the digestive tract. Kit immunohistochemistry reliably identifies the location of these cells and provides information on changes in ICC distribution and density. Human oesophagus specimens (7 embryos, 23 fetuses at 7-27 weeks gestational age; both sexes) were exposed to Kit antibodies to determine ICC differentiation. Enteric plexuses were examined immunohistochemically by using anti-neuron-specific enolase, whereas the differentiation of SMCs was studied with antibodies against α-smooth-muscle actin and desmin. By week 7, c-kit-immunopositive cells were present along the entire oesophagus in the form of an uninterrupted layer around the myenteric plexus (MP) elements. From the beginning of the 3rd month, the number of ICC progressively decreased around the MP ganglia but increased within the muscle layers. Concomitantly, differences in the number and distribution of ICC were established in the various portions of the oesophagus: specifically, ICC were abundant in the lower portion, less numerous in the middle region and rare in the upper part. By the 5th month of development, the relationship as found in later developmental stages had been established: C-kit IR ICC were present within the circular muscle layer, within the longitudinal layer and in the connective septa surrounding the muscle bundles but were completely missing around the MP ganglia.     

AC133+ progenitor cells as gene delivery vehicle and cellular probe in subcutaneous tumor models: a preliminary study

Background  

Despite enormous progress in gene therapy for breast cancer, an optimal systemic vehicle for delivering gene products to the target tissue is still lacking. The purpose of this study was to determine whether AC133+ progenitor cells (APC) can be used as both gene delivery vehicles and cellular probes for magnetic resonance imaging (MRI). In this study, we used superparamagentic iron oxide (SPIO)-labeled APCs to carry the human sodium iodide symporter (hNIS) gene to the sites of implanted breast cancer in mouse model. In vivo real time tracking of these cells was performed by MRI and expression of hNIS was determined by Tc-99m pertechnetate (Tc-99m) scan.