Solochrome Cyanine Research Articles

Schwann Cell Remyelination in the Multiple Sclerosis Central Nervous System

Multiple sclerosis (MS) is a central nervous system (CNS) demyelinating disease. Failure to remyelinate successfully is common in MS lesions, often with consequent neuronal/axonal damage. CNS myelin is normally produced by oligodendroglial cells. Remyelination by Schwann cells (SchC) has been reported in spinal cord demyelination, in which SchCs are in close proximity to CNS myelin. We identified an MS cerebral lesion that was remyelinated by SchCs. This prompted us to query the extent of SchC remyelination in the brain and spinal cords of additional autopsied MS specimens. CNS tissues were obtained from the autopsies of 14 MS cases. Remyelinated lesions were identified by Luxol fast blue-periodic-acid Schiff and solochrome cyanine staining. Deparaffinized sections containing remyelinated lesions were stained with anti–glial fibrillary acid protein to identify reactive astrocytes. Glycoprotein P zero (P0) is a protein exclusive to peripheral but not CNS myelin. Areas of SchC remyelination were identified by staining with anti-P0. Myelinated regions in the index case cerebral lesion were confirmed to be of SchC origin using anti-P0 staining. Subsequently, 64 MS lesions from 14 autopsied MS cases were examined, and 23 lesions in 6 cases showed remyelination by SchCs. Lesions from the cerebrum, brainstem, and spinal cord were examined in each case. When present, SchC remyelination was most commonly located adjacent to the venules and associated with a lower surrounding density of glial fibrillary acid protein+ reactive astrocytes than areas of only oligodendroglial cell remyelination. The difference was significant only for spinal cord and brainstem lesions but not for lesions located in the brain. In conclusion, we demonstrated SchC remyelination in the cerebrum, brainstem, and spinal cord of 6 autopsied MS cases. To our knowledge, this is the first report of supratentorial SchC remyelination in MS.

Solvothermal fabrication of different morphologies of iron doped vanadium sulfide for photocatalytic applications

This article presents the synthesis of different morphologies of iron doped vanadium sulfide by the reaction of vanadium trichloride and sodium sulfide in bare iron autoclave at 140 °C for 4 hrs using ethanol as a solvent and hydrazine as a reducing agent. S-sulfosalicylic acid was used in 1:1, 1:2 and 1:3 with vanadium trichloride keeping all the other conditions same to study the changes in morphologies of the synthesized products. Products were characterized with PXRD, SEM-EDS, ICP-OES, FTIR, TGA and UV–vis spectroscopy. Mechanism of morphological changes and photo-excitation of solochrome cyanine R under direct sunlight was evaluated in detail with UV–vis spectroscopy.

Certification procedures used by the Biological Stain Commission for eriochrome cyanine R (C.I. 43820, Mordant blue 3)

ABSTRACTEriochrome cyanine R (C.I. 43820, Mordant blue 3), also known as chromoxane cyanine R and solochrome cyanine R, has been used as a biological stain since 1957. In conjunction with ferric ions, it provides selective blue coloration of the nuclei of cells in methods procedurally similar to commonly used progressive or regressive hemalum (aluminum-hematoxylin) stains. Eriochrome cyanine R also is used to stain the myelin sheaths of axons in nerve tissue; the results are visually similar to those in sections stained with luxol fast blue MBS (C.I. 74180, solvent blue 38) with selective blue coloration of myelin and erythrocytes. Eriochrome cyanine R is an article of commerce with many uses in industrial coloration and analytical chemistry; it can be used instead of either hematoxylin or luxol fast blue MBS, especially in the event of a shortage of either of the latter compounds. The Biological Stain Commission (BSC) will certify batches of eriochrome cyanine R that meet the criteria set out in this document. The criteria include satisfactory UV/visible spectra at pH 4 and pH 12 − 13, a dye content not less than 40% and not greater than 52% (calculated as the color acid; equivalent to 46 − 59% of the trisodium salt), and satisfactory performance in three staining methods: regressive for nuclei, progressive for nuclei and regressive for myelin.

Kinetic, Isothermal and Thermodynamic Investigations for the Use of Pumpkin Seed Husks as low-Cost Biosorbent of Solochrome Cyanine R from Aqueous Solutions

Pumpkin seed husks were used in this research as effective-cost material for the adsorption of Solochrome cyanine R from aqueous solutions. A good removal ability of Pumpkin seed husks was noticed during this study, and the optimal adsorption conditions were found to be 120 min. equilibrium contact time, 0.2 g biosorbent dosage, and 100 mg/L initial dye concentration. The kinetic investigation shows that applying the pseudo-second order model gives a better fit for the adsorption with the higher correlation coefficient. The isothermal investigation shows an excellent fit of data using the Langmuir model, and also predict that the removal process is favorable. The adsorption data at different temperatures were evaluated in order to obtain the thermodynamic parameters. The positive values of ΔH° and ΔS° indicate the endothermic and randomness of the adsorption process. Moreover, the negative values of ΔG° clarify that the adsorption process is spontaneous and more preferable at higher temperatures. Pumpkin seed husks as a low-cost biosorbent exhibit a great potential for the adsorption of Solochrome cyanine R from aqueous solutions.

Solochrome cyanine: A histological stain for cobalt-chromium wear particles in metal-on-metal periprosthetic tissues

Metal-on-metal (MoM) hip arthroplasties produce abundant implant-derived wear debris composed mainly of cobalt (Co) and chromium (Cr). Cobalt-chromium (Co-Cr) wear particles are difficult to identify histologically and need to be distinguished from other wear particle types and endogenous components (e.g., haemosiderin, fibrin) which may be present in MoM periprosthetic tissues. In this study we sought to determine whether histological stains that have an affinity for metals are useful in identifying Co-Cr wear debris in MoM periprosthetic tissues. Histological sections of periprosthetic tissue from 30 failed MoM hip arthroplasties were stained with haematoxylin-eosin (HE), Solochrome Cyanine (SC), Solochrome Azurine (SA) and Perls’ Prussian Blue (PB). Sections of periprosthetic tissue from 10 cases of non-MoM arthroplasties using other implant biomaterials, including titanium, ceramic, polymethylmethacrylate (PMMA) and ultra-high molecular weight polyethylene (UHMWP) were similarly analysed. Sections of 10 cases of haemosiderin-containing knee tenosynovial giant cell tumour (TSGCT) were also stained with HE, SC, SA and PB. In MoM periprosthetic tissues, SC stained metal debris in phagocytic macrophages and in the superficial necrotic zone which exhibited little or no trichrome staining for fibrin. In non-MoM periprosthetic tissues, UHMWP, PMMA, ceramic and titanium particles were not stained by SC. Prussian Blue, but not SC or SA, stained haemosiderin deposits in MoM periprosthetic tissues and TSGT. Our findings show that SC staining (most likely Cr-associated) is useful in distinguishing Co-Cr wear particles from other metal/non-metal wear particles types in histological preparations of periprosthetic tissue and that SC reliably distinguishes haemosiderin from Co-Cr wear debris.

Environmental enrichment reduces brain damage in hydrocephalic immature rats.

We investigate the effects of environmental enrichment (EE) on morphological alterations in different brain structures of pup rats submitted to hydrocephalus condition. Hydrocephalus was induced in 7-day-old pup rats by injection of 20% kaolin into the cisterna magna. Ventricular dilatation and magnetization transfer to analyze myelin were assessed by magnetic resonance. Hydrocephalic and control rats exposed to EE (n=10 per group) were housed in cages with a tunnel, ramp, and colored plastic balls that would emit sound when touched. The walls of the housing were decorated with colored adhesive tape. Moreover, tactile and auditory stimulation was performed daily throughout the experiment. Hydrocephalic and control rats not exposed to EE (n=10 per group) were allocated singly in standard cages. All animals were weighed daily and exposed to open-field conditions every 2days until the end of the experiment when they were sacrificed and the brains removed for histology and immunohistochemistry. Solochrome cyanine staining was performed to assess the thickness of the corpus callosum. The glial fibrillary acidic protein method was used to evaluate reactive astrocytes, and the Ki67 method to assess cellular proliferation in the subventricular zone. The hydrocephalic animals exposed to EE showed better performance in Open Field tests (p<0.05), while presenting lower weight gain. In addition, these animals showed better myelination as revealed by magnetization transfer (p<0.05). Finally, the EE group showed a reduction in reactive astrocytes by means of glial fibrillary acidic protein immunostaining and preservation of the proliferation potential of progenitor cells. The results suggest that EE can protect the developing brain against damaging effects caused by hydrocephalus.

Immune characteristics study of AG490, a signal pathway inhibitor, in EAE model mice.

Multiple sclerosis (MS) is an autoimmune disease characterized by demyelination, axonal damage and progressive neurologic dysfunction in central nervous system (CNS). Many evidences show that B cells play an important role in the pathogenesis of MS. Follicular helper T cells (Tfh) secrete IL-21 to prompt the proliferation and differentiation of B cells in germinal center (GC) through clonal proliferation, somatic hypermutation, antibody class switching, antibody affinity maturation process. AG490 is a synthetic inhibitor to JAK-STAT signal pathway, which has been studied in inflammatory, tumor and autoimmune diseases. In the present study, the experimental mice were divided into 3 groups, vehicle group and AG490 group were given MOG35-55 to induce EAE model, from the third day after immunization, the mice were given vehicle or AG490 by intraperitoneal injection every other day. All mice were assessed clinical scores after immunization. On twentieth day, all mice were sacrificed, HE staining and solochrome cyanine staining were performed to evaluate inflammatory cells infiltration and demyelination, spleen sections were stained with PNA-FITC to analyze the difference in germinal center. Compared with vehicle group, the incidence of AG490 group was deceased, onset time was delayed, the severity was significantly reduced. The inflammatory cells and demyelination in AG490 group were lower than those in vehicle group. Immunofluorescence showed the fluorescence intensity of AG490 group was significantly lower than in the vehicle group, but higher than that of control group.

Pre- and Postshunting Magnetization Transfer Ratios Are in Accordance with Neurological and Behavioral Changes in Hydrocephalic Immature Rats

Hydrocephalus is a common neurological condition in children characterized by an imbalance between the production and absorption of cerebrospinal fluid (CSF), causing abnormal fluid accumulation in the brain cavities. Shunt systems have been used to drain excess CSF and to prevent progressive ventricular enlargement. However, despite improvements in these systems, neurological and structural changes cannot always be reversed. Our aim was to evaluate the magnetization transfer ratio as a biomarker for the effectiveness of a CSF shunt system to treat neurological and behavioral disorders observed in experimental hydrocephalus. Seven-day-old Wistar rats were used in this study. The pups were subjected to hydrocephalus induction via 20% kaolin intracisternal injection. After confirmation of ventriculomegaly by magnetic resonance imaging (MRI), a group of animals underwent placement of a ventriculosubcutaneous shunt (VSS). The reduction in ventricular size in hydrocephalic rats operated with functional VSS was observed as a decrease in ventricular ratio values and preservation of the corpus callosum thickness. Magnetization transfer values were significantly increased and matched to the recovery process of axonal myelination observed based on more-intense blue staining by solochrome cyanin. The histopathological analysis revealed a reduction in reactive astrocytes by means of GFAP immunostaining. The hydrocephalic rats operated with functional VSS also showed significant progress in motor and exploratory activities, similar to the control animals, at the end of the experiment. In conclusion, the VSS system employed 7 days after hydrocephalus induction was able to prevent structural damage and restore the axonal myelination process in periventricular structures by stabilizing and reducing the ventricular enlargement, and the results are in accordance with the magnetization transfer ratio in MRI.

Damage to the optic chiasm in myelin oligodendrocyte glycoprotein-experimental autoimmune encephalomyelitis mice.

Optic chiasm lesions in myelin oligodendrocyte glycoprotein (MOG)–experimental autoimmune encephalomyelitis (EAE) mice were characterized using magnetic resonance imaging (MRI) and validated using electron microscopy (EM). MR images were collected from 3 days after induction to remission, approximately 20 days after induction. Hematoxylin and eosin, solochrome cyanin–stained sections, and EM images were obtained from the optic chiasms of some mice approximately 4 days after disease onset when their scores were thought to be the highest. T2-weighted imaging and apparent diffusion coefficient map hyperintensities corresponded to abnormalities in the optic chiasms of EAE mice. Mixed inflammation was concentrated at the lateral surface. Degeneration of oligodendrocytes, myelin, and early axonal damage were also apparent. A marked increase in chiasm thickness was observed. T2-weighted and diffusion-weighted MRI can detect abnormalities in the optic chiasms of MOG-EAE mice. MRI is an important method in the study of this model toward understanding optic neuritis.

Combined therapy with methylprednisolone and ulinastatin in experimental autoimmune encephalomyelitis

Background Our previous study had demonstrated that ulinastatin (UTI) had a neuroprotective effect in experimental autoimmune encephalomyelitis (EAE). Methylprednisolone has been recommended to be a standard drug in multiple sclerosis (MS) therapies. The present study was to investigate the protective effects of UTI combined methylprednisolone in EAE. Methods Mice were divided into a UTI treatment group, a methylprednisolone treatment group, a combined treatment group with UTI and methylprednisolone, a normal saline treatment group, and a normal control group. EAE mice were induced in groups receiving different combined treatments, or respective monotherapies. Demyelination was evaluated by Solochrome cyanin staining. 2′,3′-cyclic nucleotide 3′- phosphodiesterase (CNP)/ myelin basic protein (MBP)/ the precursor form of nerve growth factor (proNGF)/p75/ inducible nitric oxide synthase (iNOS) proteins in cerebral cortex of EAE were detected by Western blotting. Results The combined treatment group had a lower clinical score ( 0.61±0.06) and demyelinating score (1.33±0.33) than the groups with normal saline (clinical score: 1.39±0.08, P &lt;0.001; demyelinating score: 2.75±0.49, P &lt;0.05) or monotheraphies. Compared with the saline treated EAE group, UTI combined methylprednisolone significantly increased expressions of CNP (1.14±0.06 vs. 0.65±0.04, P &lt;0.001), MBP (1.28±0.14 vs. 0.44±0.17, P &lt;0.001), and decreased expressions of proNGF (1.08±0.10 vs. 2.32±0.12, P &lt;0.001), p75 (1.13±0.13 vs. 2.33±0.17, P &lt;0.001), and iNOS (1.05±0.31 vs. 2.17±0.13, P &lt;0.001) proteins in EAE. Furthermore, UTI combined methylprednisolone could significantly upregulate MBP (1.28±0.14 vs. 1.01±0.15, P &lt;0.05) expression and downregulate iNOS (1.05±0.31 vs. 1.35±0.14, P &lt;0.05) expression compared to methylprednisolone treatment EAE group. And proNGF expression was significantly lower in combined treatment (1.08±0.10) than that in UTI (1.51±0.24, P &lt;0.05) or methylprednisolone (1.31±0.04, P &lt;0.05) treatment group. Conclusion Combination treatment of UTI with methylprednisolone was shown to protect EAE, suggesting that combination therapy is a potential novel treatment in MS.

Y-39983, a Selective Rho-Kinase Inhibitor, Attenuates Experimental Autoimmune Encephalomyelitis via Inhibition of Demyelination

Objective: Rho-associated kinase (ROCK) is a serine/threonine kinase and a major downstream effector of the small GTP-binding protein, Rho. Rho-ROCK triggers an intracellular signaling cascade that controls actin cytoskeleton and is essential for cell motility and adhesion, neurite outgrowth and retraction. In chronic disabling disease, multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE), demyelination and axonal damage are the major pathological changes contributing to neurological disability. We investigated the protective effect of a specific ROCK inhibitor, Y-39983, on demyelination and axonal damage in chronic EAE. Methods: Western blotting for myelin proteins, electron microscopy and solochrome cyanine staining was performed to evaluate demyelination while neurofilament proteins and cytoskeletal proteins including β-actin and β-tubulin were used to determine axonal damage in a chronic mouse model of EAE treated with Y-39983. Results: Y-39983 significantly suppressed clinical symptoms of EAE and prevented its relapse while increasing the amount of myelin proteins. No significant changes in neurofilaments and cytoskeletal proteins were observed compared with control EAE mice. The inhibition of demyelination by Y-39983 was confirmed by solochrome cyanine staining and electron microscopy. To further study the effect of Y-39983 on demyelination in EAE, we tested three major ROCK substrates, including myosin light chain phosphorylation, LIMK2 and collapsin response mediator protein-2. The activity of these molecules was decreased in EAE animals treated with Y-39983. Conclusion: The inhibitory effect of Y-39983 on demyelination is probably due to the inactivation of ROCK substrates, which are important for neurite outgrowth, growth cone collapse and demyelination of oligodendrocytes.

Electrochemical detection of dopamine at poly(solochrome cyanine)/Pd nanoparticles doped modified carbon paste electrode and simultaneous resolution in the presence of ascorbic acid and uric acid: a voltammetric method

A novel chemically modified electrode was constructed based on the electrodeposition of palladium nanoparticles (PdNps) on to a poly(solochrome cyanine) (polySCCy) film coated carbon paste electrode (CPE). Cyclic voltammetry (CV), differential pulse voltammetry (DPV), and electrochemical impedance spectroscopy (EIS) were used to characterize the properties of the modified electrode. The polySCCy/PdNPs/CPE was used for the impressionable, individualized determination and simultaneous resolution of dopamine (DA), ascorbic acid (AA) and uric acid (UA) in 0.1 M acetate buffer solution (ABS) of pH 5.0 by employing CV and DPV. The polySCCy/PdNPs/CPE exhibited strong electrocatalytic activity towards the oxidation of a mixture solution of DA, AA and UA with obvious redox potentials. The overlapping voltammetric response of biomolecules were separated into three well resolved oxidation peaks for DA, AA and UA with lower oxidation potentials and significant increase in the anodic peak currents in the presence of polySCCy/PdNPs/CPE by using CV and DPV. The results showed good sensitivity, selectivity and high reproducibility of the electrosynthesized polymer nanoparticles inclusion electrode. The limit of detection, limit of quantification and correlation coefficient of DA at polySCCy/PdNPs/CPE were 1.87 μM, 6.24 μM and 0.99375, respectively. The developed chemical sensor is ideal for the analysis of DA in pharmaceutical formulations providing satisfactory results. The interfacial electron transfer rate of DA was studied by EIS.

Hematoxylin shortages: their causes and duration, and other dyes that can replace hemalum in routine hematoxylin and eosin staining

The origins of repeated hematoxylin shortages are outlined. Lack of integration in the hematoxylin trade exacerbates the problems inherent in using a natural product. Separate corporations are engaged in tree growth and harvesting, dye extraction, processing of extracts to yield hematoxylin, and formulation and sale of hematoxylin staining solutions to the end users in biomedical laboratories. Hematoxylin has many uses in biological staining and no single dye can replace it for all applications. Probably, the most satisfactory substitutes for aluminum-hematoxylin (hemalum) are the ferric complexes of celestine blue (CI 51050; mordant blue 14) and eriochrome cyanine R (CI 43820; mordant blue 3, also known as chromoxane cyanine R and solochrome cyanine R). The iron-celestine blue complex is a cationic dye that binds to nucleic acids and other polyanions, such as those of cartilage matrix and mast cell granules. Complexes of iron with eriochrome cyanine R are anionic and give selective nuclear staining similar to that obtained with acidic hemalum solutions. Iron complexes of gallein (CI 45445; mordant violet 25), a hydroxyxanthene dye, can replace iron-hematoxylin in formulations for staining nuclei, myelin, and protozoa.

Effects of electro-acupuncture on remyelination of the transected sciatic nerve in rats

Objective To investigate the effects of electro-acupuncture (EA) on remyelination after sciatic nerve was transected in rats, so as to explore the possible mechanism of EA in treatment of peripheral nerve injury. Methods After transection of rat's sciatic nerve, the nerve regeneration chamber was built,then the rats were given EA stimulation at frequencies of 5 Hz and 100 Hz, respectively, and Solochrome Cyanine staining, HE staining of nerve fibers, immunohistochemical staining and semi-quantitative image analysis technique were employed to observe the remyelination of the nerve fibers and S-100 protein expression of Schwann cells. Results EA could significantly promote remyelination of the nerve fibers, elevate S-100 protein expression of Schwann Cells in repairing process of transected sciatic nerve of rats. The therapeutic effect of 5 Hz EA group was better than that of 100 Hz EA group. Conclusion EA is an effective way for the treatment of peripheral nerve injury. It can promote proliferation of Schwann cells and nerve fiber myelination. The therapeutic effects of EA are frequency-depondent. Key words: Electro-acupuncture; Peripheral nerve injury; Remyelination; S-100 protein

Overexpression of CNTF in Mesenchymal Stem Cells reduces demyelination and induces clinical recovery in experimental autoimmune encephalomyelitis mice

Human Mesenchymal Stem Cells (MSCs) were previously reported to ameliorate neuronal functional deficits in the MOG35-55-induced experimental autoimmune encephalomyelitis (EAE) mice by inducing T cell anergy. Human Ciliary neurotrophic factor (CNTF) recently was found to promote myelogenesis and reduce inflammation in CNTF-deficient EAE mice. We ectopically overexpressed CNTF in human MSCs to investigate its potential role in promoting remyelination and improving functional recovery in EAE mice. MSCs transfected by Ad-CNTF–IRES–EGFP (MSC–CNTF) were injected intravenously into EAE mice 10 days after the immunization. Neurological functional tests were scored daily by grading clinical signs (score 0–6). Immunofluorescence microscopy was used to detect MSC–CNTF in spinal cord. Expression of NG2, CNTF, and cleaved caspase-3 was measured by immunohistochemistry. CNTF expression was also analyzed by Western blot. Myelin was detected by Solochrome Cyanin staining. Our results found that CNTF concentration in MSC–CNTF cells was 20-fold higher than that in either MSC or Ad-EGFP-transfected MSCs (MSC–EGFP) in vitro. Mice receiving MSC–CNTF cells showed remarkable neuronal functional recovery: the cumulative clinical scores were significantly decreased, and the disease onset was statistically delayed. Mice receiving MSC–CNTF cells showed reduced TNF-α, IFN-γ and increased the level of cytokine IL-10 in peripheral blood and a large number of MSC–CNTF cells were detected in the spleen, but were not detected in other organs such as lung, liver and kidney. In the lesions of these mice, 1) the number of cleaved caspase3-positive cells was significantly reduced; 2) MSC–CNTF- and NG2-positive cells were significantly increased; and 3) the expression of CNTF was dramatically increased. In addition, demyelination was significantly reduced in MSC–CNTF mice. These data indicated that MSC–CNTF may improve functional recovery in EAE mice, possibly by exerting their immunoregulatory activity, inhibiting inflammation, homing MSC–CNTF cells to the lesions, elevating CNTF expression, reducing demyelination, and stimulating oligodendrogenesis.

Magnetic resonance imaging versus histological assessment for estimation of lesion volume after experimental spinal cord injury

The purpose of this study was to compare measures of lesion volume obtained by means of 1.5-T MR imaging to those obtained by the Cavalieri method, 6 weeks after experimental spinal cord injury. Nine male Wistar rats were subjected to spinal cord injury by clip compression (50 g) at the T-4 level. Six weeks postinjury, the rats were sacrificed, and spinal cords were analyzed ex vivo for lesion volume by means of 1.5-T MR imaging and subsequently, by the Cavalieri method. In the latter method, cords were cut longitudinally in 25-microm sections and stained with solochrome cyanin for myelin. The area of the lesion was determined for each serial section, and the distance-weighted sum of all area measures was then calculated to estimate the total lesion volume. Bland-Altman analysis showed that the 2 methods had an acceptable level of agreement for lesion volume estimation, but the Cavalieri method was prone to an overestimation bias. The MR imaging estimates of lesion volume were greater than the Cavalieri method estimates in 3 spinal cords, but the difference between measures was within 1 standard deviation of perfect agreement in these 3 lesions, and the mean difference between measures was 18.3%. In contrast, in those lesions in which the Cavalieri method yielded larger lesion volumes (5 lesions), the difference between measures was 2 standard deviations away from perfect agreement for 2 animals and the mean difference between measures was 72.4%. The results illustrate that the overestimation bias of the Cavalieri method is due, in part, to artifacts produced during processing of the spinal cord tissue.

Histochemistry of Staining Methods for Normal and Degenerating Myelin in the Central and Peripheral Nervous Systems

This review summarizes the function, structure, and chemistry of the myelin sheaths of axons: discusses the mechanisms of several staining techniques using inorganic compounds, dyes, and antibodies; and provides technical instructions for 13 staining methods for normal and degenerating myelin. Myelin has alternating hydrophobic and hydrophilic layers. The former contain the hydrocarbon tails of lipid molecules and hydrophobic segments of transmembrane proteins; the latter contain the anionic heads of polar lipids and also basic proteins. Aqueous fixatives allow some myelin lipids to be retained in paraffin sections. Stains for normal myelin include histochemical reactions that detect choline-containing phospholipids (Baker's chromation-acid hematein) or double bonds (unsaturation) in the fatty acid components of lipids (osmium tetroxide or palladium chloride reduction; the pseudoplamal reaction and DAB fluorescence, for oxidized double bonds). Traditional fat stains (Sudan dyes) stain normal myelin less intensely than fat or degenerated myelin. Dyes of the luxol fast blue group have colored anions balanced by hydrophobic organic cations. Applied from an organic solvent and differentiated in an aqueous alkali, these dyes are retained in myelin, which is more hydrophilic than fat but more hydrophobic than cytoplasm or collagen. Dye-metal complexes, notably those of ferric iron with hematein (“iron-hematoxylin”) or eriochrome cyanine R (also called chromoxane cyanine R, solochrome cyanine R, and Mordant blue 3), are excellent myelin stains for paraffin sections. They probably attach to basic proteins and resist extraction from a relatively hydrophobic environment. Immunohistochemical methods can detect characteristic proteins and lipids of myelin, some of which are different in the central and peripheral nervous systems.When an axon is disconnected from its neuronal cell body, or if the cell body dies, both the axon and its myelin sheath degenerate. Myelin degenerates also in autoimmune and toxic diseases. The hydrophobic products of degeneration contain crystals of cholesterol esters, brightly visible in frozen sections viewed with crossed polars. Hydrophobic lipids of degenerated and phagocytosed myelin take up fat stains (Sudan dyes, Nile red) but they are not colored by the myelin stains used on paraffin sections. Degenerating myelin fragments are selectively blackened in Marchi's reaction, which is with an aqueous solution containing osmium tetroxide and a hydrophilic oxidant (K2Cr207 or KClO3). (The J Histotechnol 30:87, 2007)Submitted March 21, 2007; accepted with revisions April 11, 2007

Effect of thermal energy produced by drilling on the facial nerve: histopathologic evaluation in guinea pigs

The effect of thermal energy due to drilling around the facial nerve canal on the facial nerve was histopathologically evaluated in four guinea pigs. The bony canal of the facial nerve was drilled using a 3 mm diamond burr for one minute. The temperature changes on the facial nerve canal were noted before and after dissection. The temporal bones of the animals were histopathologically examined under light microscopy using haematoxylin & eosin (H&E) and solochrome cyanine staining for myelin, and immunohistochemical staining for neuronal nitric oxide synthase (nNOS). Compared to the control group, it was observed with H&E staining that there was oedema among the axonal fibres and with solochrome cyanine staining that the thickness of the myelin fibres was decreased, and that the severity and extent of nNOS activity was decreased in the axonal fibres. It was concluded that a temperature increase on the facial canal may potentially lead to inflammation of the nerve, and may also cause deterioration of nerve conduction to some extent.

Bone mineralization density and femoral neck fragility

The traditional view of osteoporotic fractures is that they result from a reduction in bone mass combined with alterations in the micro-architecture. Apart from the effects of bone remodeling, the material properties of the remaining bone are thought to be unaffected. To test this, we compared the degree of matrix mineralization in femoral neck biopsies taken from cases of intracapsular hip fracture with age- and sex-matched postmortem controls. Whole femoral neck biopsies from seven female hip fracture cases (72–90 years) and nine controls (68–94 years) were embedded in methylmethacrylate, and sections stained with Solochrome Cyanin R for analysis of osteoid. The blocks were then diamond micro-milled, carbon coated, and analyzed for the degree of matrix mineralization using halogenated dimethacrylate standards for quantitative backscattered electron (qBSE) imaging (20 kV, entire block face, sampling interval 5 μm). The BSE gray scale was adjusted such that 0 corresponds to an electron backscattering coefficient of 0.1159 (approximately 1.70 g/ml) and 255–0.1519 (approximately 2.18 g/ml). Remodeling and mineralization data were analyzed for both the whole biopsy face and on a regional (anterior; inferior, posterior, or superior) basis. Over the whole biopsy, the level of mineralization was lower in the cases than the postmortem controls (−2.8%, P < 0.05). In both cases and controls, cortical mineralization was higher in the inferior (compressive) region compared with superior (tensile) region ( P < 0.05). Mineralization was lower in all regions of the cases (inferior: −3.3%; posterior: −3.1%; anterior: −2.7%; superior: −1.6%) compared to the controls. Mineralization density in cancellous bone was not regionally dependent but was lower in the fracture cases (−3.5%; P = 0.001). Although there were weak relationships between osteoid formation (%O.Ar/B.Ar) and the mean level of mineralization in both cortical ( P = 0.068) and cancellous ( P < 0.01) bone, adjustment for this did not markedly affect the case–control differences. In conclusion, this study has shown that in cases of intracapsular hip fracture, matrix mineralization is reduced in the femoral neck. Unexpectedly, in view of the likely role of mild to moderate vitamin D deficiency osteopathy in hip fracture, this decreased mineralization was independent of osteoid indices and therefore potentially independent of bone age. This raises the possibility that alterations in the bone matrix such as excessive glycation or changes in the composition of the collagen fibrils affect its mineralization in hip fracture cases.

Hepatic myospherulosis complicating portal vein embolisation

Aims: Myospherulosis is a rare condition characterised by sac-like structures containing spheroid bodies in cysts or cystic spaces in the tissue. This condition has not previously been reported in the...