Ultrastructural Studies of Crystal-Organic Matrix Relations in Renal Stones

Biochemically the organic matrix of kidney stones contains mucoproteins, mucopolysaccharides, inorganic material and bound water. Morphologically, the organic matrix exists as either amorphous or fibrous forms. We have attempted to critically evaluate results from analytical and morphological studies on stone matrices using light microscopy, histochemistry, x-ray diffraction, scanning electron microscopy, x-ray energy dispersive spectrometry, transmission electron microscopy and selected area electron diffraction. On the surface of calcium oxalate stones, there are usually large masses of randomly deposited calcium oxalate crystals each coated with organic matrix. Transmission electron microscopy shows these large surface crystals are composed of rows of smaller crystallites interleaved by organic matrix in a fairly orderly manner suggesting the crystallites are held together by organic matrix. In the core of a calcium containing stone, the organic matrix frequently exists as concentric laminations alternating as calcium apatite covered fibrous matrix layers and amorphous matrix layers. Transmission electron microscopy suggests that the fibrous area is probably just an area heavily populated by calcium apatite crystallites which give the fibrous appearance while the amorphous area is sparsely populated. Organic matrix richness in stones can be associated with infection and calcium apatite crystal deposition is favored in infection stones

Fictitious Calculi and Human Calculi with Foreign Nuclei

The correlative approach employing polarized light microscopy, x-ray powder diffraction, scanning electron microscopy, transmission electron microscopy, selected area electron diffraction and energy dispersive x-ray microanalysis proves to be very useful in identifying fictitious calculi and genuine human calculi with foreign body nuclei. The common artifacts as reported in the literature and observed also by us were minerals, vegetable and plant seeds, cereals, sand grains and sea shell fragments. Two interesting cases involving foreign body nuclei have been reported: one urinary calculus containing a piece of plastic-coated titanium foil in the center; one nasal calculus with a nut as a nucleus. Another common cause for foreign body nucleation is iatrogenic: intrauterine devices, catheters, suture materials and even surgical staples have been reported in the literature to be potent nidi for calculus formation. These cases remind us of the important fact that our body fluids are supersaturated with respect to calcium phosphates and occasionally to other compounds. Hydroxyapatite crystals are readily nucleated by foreign bodies. Whitlockite is involved if the fluid Mg/Ca ratio is in a suitable range, brushite if the fluid is acidic and struvite if there is urea-splitting infection. In urine and other fluids, calcium oxalate and uric acid crystals contribute to the calculus growth

Crystal Associated Diseases: Role of Scanning Electron Microscopy in Diagnosis

As crystals are important etiologic agents for disease, their accurate identification in tissues and body fluids is of utmost importance. This paper surveys the roles of crystals in disease process and outlines current analytical techniques for crystal detection and identification in bone tissues. The value of multiple correlated techniques is demonstrated including scanning electron microscopy, x-ray energy spectroscopy and powder diffraction analysis. The current feasibility of utilizing intermediate voltage scanning transmission analytical electron microscopy to integrate these analytical techniques on the same tissue sample is emphasized

Early stages of ramified growth in quasi-two-dimensional electrochemical deposition

I have measured the early stages of the growth of branched metal aggregates formed by electrochemical deposition in very thin layers. The growth rate of spatial Fourier modes is described qualitatively by the results of a linear stability analysis [D.P. Barkey, R.H. Muller, and C.W. Tobias, J. Electrochem. Soc. {\bf 136}, 2207 (1989)]. The maximum growth rate is proportional to $(I/c)^\delta$ where $I$ is the current through the electrochemical cell, $c$ the electrolyte concentration, and $\delta = 1.37 \pm 0.08$. Differences between my results and the theoretical predictions suggest that electroconvection in the electrolyte has a large influence on the instability leading to ramified growth.Comment: REVTeX, four ps figure

Determining collagen distribution in articular cartilage using contrast-enhanced micro-computed tomography

Objective: Collagen distribution within articular cartilage (AC) is typically evaluated from histological sections, e.g., using collagen staining and light microscopy (LM). Unfortunately, all techniques based on histological sections are time-consuming, destructive, and without extraordinary effort, limited to two dimensions. This study investigates whether phosphotungstic acid (PTA) and phosphomolybdic acid (PMA), two collagen-specific markers and X-ray absorbers, could (1) produce contrast for AC X-ray imaging or (2) be used to detect collagen distribution within AC. Method: We labeled equine AC samples with PTA or PMA and imaged them with micro-computed tomography (micro-CT) at pre-defined time points 0, 18, 36, 54, 72, 90, 180, 270 h during staining. The micro-CT image intensity was compared with collagen distributions obtained with a reference technique, i.e., Fourier-transform infrared imaging (FTIRI). The labeling time and contrast agent producing highest association (Pearson correlation, BlandeAltman analysis) between FTIRI collagen distribution and micro-CT -determined PTA distribution was selected for human AC. Results: Both, PTA and PMA labeling permitted visualization of AC features using micro-CT in non-calcified cartilage. After labeling the samples for 36 h in PTA, the spatial distribution of X-ray attenuation correlated highly with the collagen distribution determined by FTIRI in both equine (mean +/- S.D. of the Pearson correlation coefficients, r = 0.96 +/- 0.03, n = 12) and human AC (r = 0.82 +/- 0.15, n = 4). Conclusions: PTA-induced X-ray attenuation is a potential marker for non-destructive detection of AC collagen distributions in 3D. This approach opens new possibilities in development of non-destructive 3D histopathological techniques for characterization of OA. (C) 2015 The Authors. Published by Elsevier Ltd and Osteoarthritis Research Society International.Peer reviewe

3D morphometric analysis of calcified cartilage properties using micro-computed tomography

Objective: Our aim is to establish methods for quantifying morphometric properties of calcified cartilage (CC) from micro-computed tomography (mu CT). Furthermore, we evaluated the feasibility of these methods in investigating relationships between osteoarthritis (OA), tidemark surface morphology and open subchondral channels (OSCCs). Method: Samples (n = 15) used in this study were harvested from human lateral tibial plateau (n = 8). Conventional roughness and parameters assessing local 3-dimensional (3D) surface variations were used to quantify the surface morphology of the CC. Subchondral channel properties (percentage, density, size) were also calculated. As a reference, histological sections were evaluated using Histopathological osteoarthritis grading (OARSI) and thickness of CC and subchondral bone (SCB) was quantified. Results: OARSI grade correlated with a decrease in local 3D variations of the tidemark surface (amount of different surface patterns (r(s) = -0.600, P = 0.018), entropy of patterns (EP) (r(s) = -0.648, P = 0.018), homogeneity index (HI) (r(s) = 0.555, P = 0.032)) and tidemark roughness (TMR) (r(s) = -0.579, P = 0.024). Amount of different patterns (ADP) and EP associated with channel area fraction (CAF) (r(p) = 0.876, P <0.0001; r(p) = 0.665, P = 0.007, respectively) and channel density (CD) (r(p) = 0.680, P = 0.011; r(p) = 0.582, P = 0.023, respectively). TMR was associated with CAF (r(p) = 0.926, P <0.0001) and average channel size (r(p) = 0.574, P = 0.025). CC topography differed statistically significantly in early OA vs healthy samples. Conclusion: We introduced a mu-CT image method to quantify 3D CC topography and perforations through CC. CC topography was associated with OARSI grade and OSCC properties; this suggests that the established methods can detect topographical changes in tidemark and CC perforations associated with OA. (c) 2018 The Authors. Published by Elsevier Ltd on behalf of Osteoarthritis Research Society International. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).Peer reviewe

The Role of Citrullinated Proteins Suggests a Novel Mechanism in the Pathogenesis of Multiple Sclerosis

The pathogenesis of MS is unknown. In our studies, we have demonstrated an important role for citrullinated myelin basic protein (MBP). The accompanying loss of positive charge compromises the ability of MBP to interact with the lipid bilayer. The conversion of arginine to citrulline in brain is carried out by an enzyme peptidyl arginine deiminase (PAD) 2. The amount of PAD 2 in brain was increased in MS normal-appearing white matter. The mechanism responsible for this increase involved hypomethylation of the promoter region in the PAD 2 gene in MS, but no change (compared to normal) was found in thymus tissue DNA from the same MS patients. In addition, no change was observed in other neurological diseases, including Alzheimer’s, Parkinson’s, and Huntington’s. We propose that citrullinated MBP, resulting from elevated levels of PAD 2 represents an important biochemical pathway in the pathogenesis of MS

Enhanced cartilage regeneration in MIA/CD-RAP deficient mice

Melanoma inhibitory activity/cartilage-derived retinoic acid-sensitive protein (MIA/CD-RAP) is a small soluble protein secreted from chondrocytes. It was identified as the prototype of a family of extracellular proteins adopting an SH3 domain-like fold. In order to study the consequences of MIA/CD-RAP deficiency in detail we used mice with a targeted gene disruption of MIA/CD-RAP (MIA−/−) and analyzed cartilage organisation and differentiation in in vivo and in vitro models. Cartilage formation and regeneration was determined in models for osteoarthritis and fracture healing in vivo, in addition to in vitro studies using mesenchymal stem cells of MIA−/− mice. Interestingly, our data suggest enhanced chondrocytic regeneration in the MIA−/− mice, modulated by enhanced proliferation and delayed differentiation. Expression analysis of cartilage tissue derived from MIA−/− mice revealed strong downregulation of nuclear RNA-binding protein 54-kDa (p54nrb), a recently described modulator of Sox9 activity. In this study, we present p54nrb as a mediator of MIA/CD-RAP to promote chondrogenesis. Taken together, our data indicate that MIA/CD-RAP is required for differentiation in cartilage potentially by regulating signaling processes during differentiation

Articular cartilage mineralization in osteoarthritis of the hip

<p>Abstract</p> <p>Background</p> <p>The aim of this study was to examine the frequency of articular cartilage calcification in patients with end-stage hip OA. Further, its impact on the clinical situation and the OA severity are analyzed.</p> <p>Methods</p> <p>Eighty patients with OA of the hip who consecutively underwent total hip replacement were prospectively evaluated, and 10 controls were included. The patients' X-rays were analyzed for the presence of articular cartilage mineralization. A Harris Hip Score (HHS) was preoperatively calculated for every patient.</p> <p>Slab specimens from the femoral head of bone and cartilage and an additional square centimeter of articular cartilage from the main chondral defect were obtained from each patient for analysis of mineralization by digital contact radiography (DCR). Histological grading was also performed. In a subset of 20 patients, minerals were characterized with an electron microscope (FE-SEM).</p> <p>Results</p> <p>Calcifications were seen in all OA cartilage and slab specimens using DCR, while preoperative X-rays revealed calcification in only 17.5%. None of the control cartilage specimens showed mineralization. There was a highly significant inverse correlation between articular cartilage calcification and preoperative HHS. Histological OA grade correlated positively with the amount of matrix calcification. FE-SEM analysis revealed basic calcium phosphate (BCP) as the predominant mineral; CPPD crystals were found in only two patients.</p> <p>Conclusions</p> <p>Articular cartilage calcification is a common event in osteoarthritis of the hip. The amount of calcification correlates with clinical symptoms and histological OA grade.</p