Point-counting Method Research Articles

Quantitative determination of minerals and anthropogenic particles in some Polish peat occurrences using a novel SEM point-counting method

A method is proposed for determining the mineral composition of peat using scanning electron microscope. In an illustrative example, five groups of particles occurring in amounts of >0.05 % are distinguished in peat from Puścizna Mała bog in the Carpathian foreland, Poland. These are spheroidal aluminosilicate particles (SAP), feldspars, nondescript aluminosilicates (mainly clays), silica (quartz and opaline silica), and Fe(hydro)oxides. Two more site-specific groups (barite and ZnS) are distinguished in highly polluted fens (Bagno Bruch and Bagno Mikołeska) near a zinc smelter in Upper Silesia. At Bagno Bruch, peat contents of predominantly authigenic ZnS microspheroids range up to 1.1 %. SAP originating from coal-burning power stations account for maximum concentrations of <21–39 % of the inorganic fraction in the studied mires. SAP concentrations vary with depth, and mean spheroid diameters with distance from emission sources. A distinct feature of SAP is their common enrichment in Ti what questions the use of Ti as a proxy for soil dust in fly ash polluted bogs. As amounts of anthropogenic magnetic spherules, less abundant than SAP in all mires, relate to water table level position, they are unsuitable as tracers of air pollution. The proposed method is recommended for application with peats having ash contents > ~4 %.

A study of regenerative ability of liver after repetitive heat stress induced liver injury

Background: It is well known that heat stress produces liver injury. It is also known that liver regenerate after injury. This way, there should be almost no or minimal alteration in the size of the liver after regeneration. Aims & Objective: In the present study, after heat induced liver injury, the ability of liver to regenerate itself was assessed. Liver injury was assessed by biochemical [Serum Glutamic Pyruvate Transaminase (SGPT); Serum Glutamic Oxaloacetic Transaminase (SGOT); and Alkaline Phosphatase (ALKP)], morphological, and morphometric changes. On the other hand, Liver regeneration was evaluated by morphological and morphometric observations of male adult albino rats (Wistar strain). Material and Methods: The experimental animals were subjected to repetitive heat stress for 4 hours daily, at 37 ± 0.5 ˚C in a Biological Oxygen Demand (BOD) incubator (relative humidity 65 - 82%) for 2 and 5 consecutive days. Biochemical assessment was done on blood collected from left ventricle of beating heart of rats. Morphometric and morphological studies were conducted under light microscope on paraffin sections (H&E) of liver from control and experimental animals. The morphometric analysis was done by intersection – point counting method, using simple square lattice test system. Relative Liver Wet Weight of all animals was calculated. Results: Progressively degenerative changes in morphological observations (disruption of cell plates in liver lobules and Kupffer cell hyperplasia), progressively increased statistical significance of morphometric (numerical density of Kupffer cells on area - Nak), and biochemical parameters informed that increasing liver damage was present with increased repetition of heat exposures in 2 and 5 days heat exposed experimental albino rats. With this liver degeneration, Relative Liver Wet Weight of all the experimental animals should have been decreased but it was not, reiterating about well-known fact of regenerative ability of liver. Along with progressive changes of heat induced liver injury, progressively increasing regenerative changes were also evident on morphological (binucleate cells and anisocytosis) observations supported by statistically significant morphometric (volume density of hepatocytes – Vvh and numerical density of hepatocytes - Nvh) parameters in experimental animals. Conclusion: The above findings suggested that the regenerative ability of liver progressively increased with progressively increasing liver injury.

Analysis of pulmonary surfactant in rat lungs after intratracheal instillation of short and long multi-walled carbon nanotubes

Multi-walled carbon nanotubes (MWCNTs) are interesting new materials, but there is some concern about their harmfulness due to their fibrous nature. To determine the difference in the biological effects of MWCMTs by fiber length, we prepared two MWCNT samples from one bulk sample. One consisted of cut up short fibers (Short; average length = 0.94 µm) and the other was just dispersed (Long; average length = 3.4 µm). The samples were administered to male Wistar rats by intratracheal instillation at doses of 0.2 mg and 1 mg/animal (Short) and 0.2 mg and 0.6 mg/animal (Long). The animals were sacrificed at time points from 3 d to 12 months after administration. Bronchoalveolar lavage fluid (BALF) was taken from the lungs and pathological specimens were prepared. The concentrations of phospholipids, total protein and surfactant protein D (SP-D) in the pulmonary surfactant of the BALF were determined, the surface tension of BALF was measured, and the inflammation score was determined by the point-counting method to assess pulmonary tissue inflammation. The present study suggests that inflammatory response in the lung was slightly higher for long MWCNTs than for short MWCNTs when compared at the same mass dose. The correlation between pulmonary surfactant components and BALF surface tension was also evaluated. The Spearman’s rank correlation coefficients obtained for the phospholipid, total protein and SP-D concentrations were −0.068 (p = 0.605), −0.360 (p = 0.005) and −0.673 (p = 0.000), respectively. Surface tension, measured by a simple method, should be reflected in the change of a surfactant protein, such as SP-D.

Detrital Mode and Geochemistry of the Shurijeh Formation (Late Jurassic‐Early Cretaceous) in the Central and Western Parts of the Intracontinental Kopet‐Dagh Basin, NE Iran: Implications for Provenance, Tectonic Setting and Weathering Processes

Abstract:Petrographical and geochemical methods were combined to investigate the provenance, geodynamic and weathering history of the Shurijeh sandstones, Kopet‐Dagh Basin. The pointcounting method and XRF technique are used for modal and geochemical analyses. Based on petrographical examinations, it seems that the Shurijeh sandstones are mainly deposited in the craton interior and recycled orogen belts. In addition to petrographical investigation, geochemical analyses (major oxides and trace elements) of Late Jurassic‐Early Cretaceous rocks reveal that the sedimentation processes are performed in a passive continental margin. Such interpretation is supported with geodynamic and paleogeographical studies of the Kopeh‐Dagh basin during this time. The geochemical investigations suggested that the composition of probable source rocks mostly was acidic‐intermediate with minor mafic igneous rocks. Based on the above, Paleo‐Tethys remnants and their collision‐related granitoids, in the south and west of Mashhad, may have been the source area for these rocks. CIA values, which range from 63.8 to 94.9 in samples, are suggesting a moderate to relatively high degree of alteration (weathering) in the source area. Therefore, petrographical and paleogeographical studies of siliciclastic rocks can be used for the provenance, tectonic setting and paleoweathering studies in the source area.

Function points as a universal software metric

Function point metrics are the most accurate and effective metrics yet developed for software sizing and also for studying software productivity, quality, costs, risks, and economic value. Unlike the older "lines of code" metric function points can be used to study requirements, design, and in fact all software activities from development through maintenance. In the future function point metrics can easily become a universal metric used for all software applications and for all software contracts in all countries. The government of Brazil already requires function points for all software contracts, and South Korea and Italy may soon follow. However, there are some logistical problems with function point metrics that need to be understood and overcome in order for function point metrics to become the primary metric for software economic analysis. Manual function point counting is too slow and costly to be used on large software projects above 10,000 function points in size. Also, application size is not constant but grows at about 2% per calendar month during development and 8% or more per calendar year for as long as software is in active use. This paper discusses a method of high-speed function point counting that can size any application in less than two minutes, and which can predict application growth during development and for five years after release. This new method is based on pattern matching and is covered by U.S. utility patent application and hence is patent pending.

A possible role for CD8+ and non‐CD8+ cell granzyme B in early small airway wall remodelling in centrilobular emphysema

CD8(+) cell infiltration and apoptosis of airway epithelial cells are increased in chronic obstructive pulmonary disease (COPD). CD8(+) T cells induce apoptosis by releasing granzymes, which can also cause extracellular matrix degradation and remodelling. Granzyme B levels and T cells expressing granzyme B are increased in bronchoalveolar lavage fluid of COPD patients, which suggests that granzyme B may contribute to the pathogenesis of COPD. This study provides quantitation of granzyme B-positive cells in relation to CD8(+) cells in the small airway walls of emphysema. Antibodies against CD8 and granzyme B were used to identify CD8(+) and granzyme B(+) cells. Volume fraction (Vv) of CD8(+) and granzyme B(+) cells were quantitated by the point counting method in the small airways of 13 non-smoker control subjects and 46 emphysema patients (14 panlobular emphysema (PLE) and 32 centrilobular emphysema (CLE) lungs). Immunohistochemical detection of macrophage scavenger receptor was also performed in randomly selected cases. The Vv of CD8(+) and granzyme B(+) cells in CLE was greater than those in control and PLE (both P < 0.001) subjects. The Vv of granzyme B(+) cells was greater than that of CD8(+) cells (P = 0.006), and not all CD8(+) cells were positive for granzyme B in CLE subjects. Monocytes expressing both granzyme B and macrophage scavenger receptor and granulocytes expressing granzyme B were identified. Upregulation of granzyme B in CD8(+) and non-CD8(+) cells is an early phenomenon of small airway wall remodelling in centrilobular emphysema and suggests its possible role in the pathogenesis of COPD.

Assessment of liver damage in male albino rats after repetitive heat stress of moderate level

Background: There are multiple studies showing the effect of severe heat stress on systemic functions. Severe heat stress has been known to affect almost every organ in the body. It is well proved that liver is the organ which is severely affected by heat stress and the biochemical, morphological, and morphometric changes of liver following severe heat stress are well documented. But, the effect of repetitive heat stress of moderate level on the liver has not been studied extensively. Aims & Objective: In the present study, the effect of repetitive heat stress of moderate level was assessed on biochemical [Serum Glutamic Pyruvate Transaminase (SGPT); Serum Glutamic Oxaloacetic Transaminase (SGOT); and Alkaline phosphatase (ALKP)], morphological, and morphometric changes of liver of adult albino male rats (wistar strain). Materials and Methods: The experimental animals were subjected to repetitive heat stress for 4 hours daily, at 37 ± 0.5oC in a Biological Oxygen Demand (BOD) incubator (relative humidity 65 - 82%) for 2, 5, and 10 consecutive days. Biochemical assessment (SGPT, SGOT, and ALKP) was done on blood collected from left ventricle of beating heart of rats. Morphometric and morphological studies were conducted under light microscope on paraffin sections (H&E) of liver from control and experimental animals. The morphometric analysis was done by intersection – point counting method, using simple square lattice test system. Results: The serum levels of liver enzymes were elevated in all heat exposed animals (statistically significant in five and ten days exposed animals) in comparison to controls. Morphological changes of anisocytosis at some sites, disruption of cell plates in lobules, mild Kupffer cell hyperplasia were present in rats exposed to heat for two consecutive days. After five days heat exposure ballooning degeneration, single cell necrosis along with small foci of necrosis disrupting cell plates in lobules, and sinusoidal compression were noticed. Kupffer cell hyperplasia was observed. At various sites, hepatocytes showed regenerative changes as binucleate cells and anisocytosis. After 10 days exposure, changes became more marked. The volume density of hepatocytes (Vvh) and numerical density of hepatocytes (Nvh) increased with the increase in heat exposure, despite increasing degenerative changes, confirming regenerative power of liver by hepatocyte proliferation. The increasing numerical density of Kupffers cells on area (Nak) indicated progressive liver damage. Conclusion: All the observations confirmed that exposure to repetitive heat stress, even of moderate level, leads to liver damage.

Defining the percentage of intra-abdominal hemorrhage in abdominal computerized tomography using stereology in patients with blunt liver injury and determining its relationship with outcomes

The aim of this study is to determine the percentage of intra-abdominal hemorrhage (PIAH) on computerized tomographic (CT) scan via the Cavalieri method and to define whether this is correlated with the outcome. Fifty-one patients (24 children and 27 adults) with blunt hepatic injury were studied in this Level III prognostic clinical cohort study. The stereologic method of point counting based on the Cavalieri approach was adapted to CT data so as to assess IAH and abdominal volume. PIAH was calculated as intra-abdominal fluid volume/whole abdominal volume × 100. Mean PIAHs in children and adults were 4.20% ± 2.85% and 6.28% ± 5.21%, respectively. Coexisting intra-abdominal injuries in children and in adults were as follows: splenic injury (29.2, 11.1%; p = 0.012), kidney (25, 11.1%), bladder (4.2, 14.8%), and pelvic fracture (12.5, 11.1%). PIAH was moderately negatively associated with hemoglobin levels (r = -0.301; p = 0.032), hematocrit levels (r = -0.322; p = 0.021), and GCS (Glasgow Coma Scale) score (r = -0.276; p = 0.05). Neither receiver operating characteristic curve analyses for PIAH nor outcomes were statistically significant in children. In adults, sensitivity and specificity of PIAH in predicting the prognoses when the cutoff levels were taken as 5.39%, 9.9%, and 12.4%, respectively, were as follows: operation (71, 84%), mortality (36, 93%), and intensive care unit admission (25, 94%). In patients with blunt hepatic injury, the Cavalieri principle of stereology can easily be added to the CT slices to calculate PIAH. This method is repeatable in other institutions and can be used as a guide to predict outcomes. It is suitable for a universal parameter to measure intra-abdominal fluid in blunt injury. PIAH has low sensitivity but high specificity to predict intensive care unit admission and mortality in cases of blunt hepatic injury in adults. Its specificity in predicting the need for operation is better than that of the anatomic liver injury grading systems in computerized tomography. Prognostic study, level III.

Comparison of two methods for the estimation of subcortical volume and asymmetry using magnetic resonance imaging: a methodological study

The subcortical brain structures are associated with other structures of nervous system; therefore, they have major influence on sensory-motor, limbic and cognitive information processing. Magnetic resonance imaging provides a detailed knowledge of normal and diseased anatomical structures for medical research. The aim of the current study was to compare the volumes of subcortical brain structures and determine the probable volumetric asymmetry in healthy subjects using stereological (point-counting) and semi-automatic segmentation methods. MR scans were obtained from 30 subjects (17 males, 13 females) free of any psychiatric, neurological or cognitive impairment. MR images were analyzed by using stereological (point-counting) and semi-automatic segmentation methods. We did not find any significant differences among the subjects with respect to gender using both methods. This study showed no significant asymmetry in subcortical structures according to methods. Also, no significant difference was found between point-counting and semi-automated segmentation methods for the volumes of subcortical structures (p>0.05). From these results, it can be concluded that the semi-automated segmentation method and stereological technique can be used for reliable volume estimation of subcortical structures. However, the stereological method takes less time than semi-automated segmentation; it is simple, reliable and inexpensive. Further studies are required with larger samples in order to support these data.

Desnutrição materna durante a lactação em ratos Wistar: efeitos sobre as fibras elásticas da matriz extracelular na traqueia dos filhotes

To investigate the effects of maternal protein malnutrition during lactation on the elastic fibers in the tracheas of Wistar rat pups. At delivery, 12 male pups of two Wistar rat dams were equally divided into two groups: control, in which the dam received water and standard rat chow ad libitum during lactation; and protein-restricted (PR), in which the dam received water ad libitum and an isoenergetic PR diet (8% protein). At 21 days of age, the pups were killed and their tracheas were excised. The elastic fibers were stained with Weigert's resorcin-fuchsin (after oxidation) and evaluated under light microscopy. Morphometric determinations were performed by stereology, with the point-counting method, and expressed as volumetric densities. Elastic fibers, most having a longitudinal distribution, were identified beneath the tracheal mucosa. In addition, well-defined circular layers of elastic fibers were found around the inner and outer surfaces of the cartilaginous ring. There were no differences between the groups regarding the organization and distribution of the elastic fibers. The volumetric density of the elastic fibers of the pups in the control and PR groups was 2.46 ± 0.99% and 3.25 ± 1.13%, respectively (p < 0.01). The volumetric density of elastic fibers appears to be greater in rat pups breastfed by dams receiving a PR diet than in those breastfed by dams receiving a normal diet.

Cleaning and modification of intraorally contaminated titanium discs with calcium phosphate powder abrasive treatment

The aim of this study was to evaluate the cleaning efficiency on intraorally contaminated titanium discs by using calcium phosphate and air powder abrasive (APA) treatment. The modification of titanium surface (SLA) was evaluated and compared with the conventional air powder abrasive methods and phosphoric acid. This treatment modality might give new perspectives for peri-implant surface treatment. A total of 36 SLA surface titanium discs were kept in the human mouth for 48 h by 14 volunteers. The intraorally contaminated discs were stained with erythrosine dye to make the biofilm visible. Discs were randomly assigned to one of the six groups: APA without powder-only water and air (Control). APA with Hydroxylapatite (HA). APA with Hydroxylapatite and Calcium Phosphate (HA + TCP). APA with Titanium Dioxide (TiO2). APA with EMS Soft Subgingival powder (EMS). Phosphoric Acid. Light microscope photos were taken during the treatment. Following the cleaning, the residual biofilm, surface changes, and surface chemical content were evaluated using Scanning Electron Microscopy (SEM) and Energy Dispersive X-Ray Spectroscopy (EDS). A systematic random sampling protocol and a point counting method were applied for the quantitative evaluation of the remaining biofilm. Multiple comparisons within and between groups are performed by Kruskall Wallis test and if significant Mann-Whitney U-test as post hoc testing is applied. The significance level was P < 0.05. All methods with the exception of phosphoric acid could decrease the initial amount of biofilm significantly. Among all air powder abrasive treatments, the HA + TCP group showed the best results with 99% biofilm removal, followed by HA and EMS powders. The cleaning method caused minimal changes to the surface structure. With the exception of the control group, all air powder applications caused sharp edges around the grooves in the implant surface to be rounded. TiO2 powder caused less change than HA and HA + TCP. Phosphoric acid did not cause a visible surface change on the SEM photos. Powder particles remnants were observed on and impacted in the titanium surface. In the HA and HA + TCP group, a Ca content was observed varying between 2% and 5%. In the control group, saliva and biofilm-related elements were observed. Using the air powder abrasive method with calcium phosphate powders on contaminated titanium discs, an efficient implant cleaning and surface modification can be achieved. This method should be further improved as it has possible potential to be used as an implant surface treatment method for implants involved with peri-implantitis.

Adipocytes derived fibrinolytic components in peritoneum — a pilot study

Abstract The proteins of the fibrinolytic system — urokinase plasminogen activator(uPA), tissue plasminogen activator (tPA)and plasminogen activator inhibitor type IPAI-I) — play important roles in fibrotization in various organs and including peritoneum. To study the cellular localization of PAI-1, tPA and uPA within the adipose tissue of the peritoneal membrane in patients at the onset of peritoneal dialysis(PD) we determined the initial expression of these proteins in relationship to multiple clinical variables. Methods: routinely performed parietal peritoneal biopsies in 12 patients undergoing peritoneal catheter implantation were examined. We used formalinfixed, paraffin-embedded specimens for immunohistochemical localization of these proteins along with the stereological pointcounting method for quantification of their expression within the peritoneal adipose tissue. Results: strong positive mutual correlation between the expression of PAI-1 and both uPA (SpearmanR=0.66) and tPA (R=0.59) as well as between the expression of uPA and tPA (R=0.77) was found without any relatioship to BMI, age, peritoneal transport characteristic or diabetes status. Conclusion: Adipose tissue within the peritoneum is capable of producing fibrinolysis regulators (independently on clinical parameters) thus possibly affecting the fibrotization and function of peritoneum as dialysis membrane. The effect of dialysis solution dosing, composition and other dialysis related factors should be clarified in future studies.

A simple stereological method for estimating the number and the volume of the pancreatic beta cells.

Number of the beta cells as well as their volume is a fundamental variable in the pancreatic research.This study describes a simple method for stereological estimation of the volume of the pancreas, the total volume of the islets, and the total number, as well as the mean volume, of the beta cells in rat.The primary volume of the pancreas was measured using the immersion method. Also, tissue shrinkage was estimated and the final pancreas volume was corrected without the need for serial sectioning. A limited number (i.e., 10-13) of the isotropic uniform random slabs of each pancreas was embedded in the same block. One 5 µm and one 20 µm sections were obtained and stained with a modified aldehyde fuchsin. The point counting, optical disector and point-sampled intercept methods were used to estimate the volume density of the islet, the numerical density of the beta cells, and the mean cell volume, respectively.After calculating the tissue shrinkage, the mean primary volume of the pancreas (628 mm3; CV: 25%) was corrected to obtain the final volume (442 mm3; CV: 39%). The mean islet volume was reported as 3.8 mm3 (CV: 22%). Besides, the total number of the beta cells was estimated as 2.9x10(6) (CV: 20%). Moreover, the mean volume of the beta cells was obtained (1,158 µm3; CV: 9%).It takes almost one hour to estimate the volume of the islets and two hours to count the cells and estimate the intercepts per animal.

Abstract 540: Plasma Cell Infiltration and Mucoid Degeneration in the Media of Ascending Aorta in Patients with Coronary Artery Disease

Introduction Atherosclerosis results in inflammatory changes in the aortic intima, but little is known regarding medial changes. The inflammatory diseases of the aorta and large artery display characteristic features of inflammatory cell infiltration in the media, and they are usually diagnosed by medial changes. Aims Atherosclerosis of the ascending aorta coexists with coronary artery disease. The aim of this study was to investigate the atherosclerotic changes in 44 biopsy specimens of media of the ascending aorta associated with coronary artery disease. Methods We compared plasma cells, and matrix metalloproteinase (MMP)-2-, -9- and -12-positive cells immunohistochemically, and we also compared mucoid degeneration and fibrosis determined by special staining using a point-counting method, for groups with a variable number of coronary stenotic (≥75 %) lesions. Results and Conclusion To investigate the appearance of plasma cells, smooth muscle cells, and mucoid degeneration in the media of the atherosclerotic aorta, we examined three cadavers with known atherosclerosis. CD38-positive plasma cells are pleomorphous in the aortic media. Desmin-positive smooth muscle cells appeared fibrous or solitary spindle cells. Mucoid degeneration was shown in all three cadaver cases. In patients with one to three coronary stenotic lesions, plasma cells and mucoid degeneration were low in the aortic media. With four to five lesions, both plasma cells and mucoid degeneration increased significantly compared with those in the group with one to three lesions, and MMP-12-positive cells significantly decreased. In patients with six to nine lesions, the number of plasma cells was significantly lower than in patients with four or five lesions, whereas mucoid degeneration significantly increased. There was no change in fibrosis. In conclusion, both the infiltration of short lived plasma cells and the exudation of proteoglycan are known to be regarded as characteristic features in the autoimmune disease. The delayed stage in our study is suggested to be caused by autoimmune reaction. If the cause of the delayed medial atherosclerosis stage is found, then more effective therapies to suppress its development can be introduced.

Stereological evaluation of the volume and volume fraction of newborns’ brain compartment and brain in magnetic resonance images

Brain development in early life is thought to be critical period in neurodevelopmental disorder. Knowledge relating to this period is currently quite limited. This study aimed to evaluate the volume relation of total brain (TB), cerebrum, cerebellum and bulbus+pons by the use of Archimedes' principle and stereological (point-counting) method and after that to compare these approaches with each other in newborns. This study was carried out on five newborn cadavers mean weighing 2.220 ± 1.056 g with no signs of neuropathology. The mean (±SD) age of the subjects was 39.7 (±1.5) weeks. The volume and volume fraction of the total brain, cerebrum, cerebellum and bulbus+pons were determined on magnetic resonance (MR) images using the point-counting approach of stereological methods and by the use of fluid displacement technique. The mean (±SD) TB, cerebrum, cerebellum and bulbus+pons volumes by fluid displacement were 271.48 ± 78.3, 256.6 ± 71.8, 12.16 ± 6.1 and 2.72 ± 1.6 cm3, respectively. By the Cavalieri principle (point-counting) using sagittal MRIs, they were 262.01 ± 74.9, 248.11 ± 68.03, 11.68 ± 6.1 and 2.21 ± 1.13 cm3, respectively. The mean (± SD) volumes by point-counting technique using axial MR images were 288.06 ± 88.5, 275.2 ± 83.1, 19.75 ± 5.3 and 2.11 ± 0.7 cm3, respectively. There were no differences between the fluid displacement and point-counting (using axial and sagittal images) for all structures (p > 0.05). This study presents the basic data for studies relative to newborn's brain volume fractions according to two methods. Stereological (point-counting) estimation may be accepted a beneficial and new tool for neurological evaluation in vivo research of the brain. Based on these techniques we introduce here, the clinician may evaluate the growth of the brain in a more efficient and precise manner.

Assessment methods for the evaluation of vitiligo

There is no standardized method for assessing vitiligo. In this article, we review the literature from 1981 to 2011 on different vitiligo assessment methods. We aim to classify the techniques available for vitiligo assessment as subjective, semi-objective or objective; microscopic or macroscopic; and as based on morphometry or colorimetry. Macroscopic morphological measurements include visual assessment, photography in natural or ultraviolet light, photography with computerized image analysis and tristimulus colorimetry or spectrophotometry. Non-invasive micromorphological methods include confocal laser microscopy (CLM). Subjective methods include clinical evaluation by a dermatologist and a vitiligo disease activity score. Semi-objective methods include the Vitiligo Area Scoring Index (VASI) and point-counting methods. Objective methods include software-based image analysis, tristimulus colorimetry, spectrophotometry and CLM. Morphometry is the measurement of the vitiliginous surface area, whereas colorimetry quantitatively analyses skin colour changes caused by erythema or pigment. Most methods involve morphometry, except for the chromameter method, which assesses colorimetry. Some image analysis software programs can assess both morphometry and colorimetry. The details of these programs (Corel Draw, Image Pro Plus, AutoCad and Photoshop) are discussed in the review. Reflectance confocal microscopy provides real-time images and has great potential for the non-invasive assessment of pigmentary lesions. In conclusion, there is no single best method for assessing vitiligo. This review revealed that VASI, the rule of nine and Wood's lamp are likely to be the best techniques available for assessing the degree of pigmentary lesions and measuring the extent and progression of vitiligo in the clinic and in clinical trials.

Age-dependent ultrastructural changes of coronary artery in spontaneously hypertensive rats

The age-dependent differences in basic cardiovascular parameters, geometry and structure of coronary arteries between Wistar and spontaneously hypertensive rats (SHR) were evaluated. SHR of the age 3-, 9-, 17-, and 52-week and age-matched Wistar rats were used. Blood pressure (BP) was measured by the plethysmographic method. Animals were perfused with a glutaraldehyde fixative under pressure of 90 mmHg (3-week-old) and 120 mmHg (9-, 17-, 52-week-old). Coronary arteries were processed for electron microscopy. The proportions and cross sectional areas (CSA) of extracellular matrix in intima and media, endothelial and muscle cells were determined by point counting method. Cardiac hypertrophy and except of 3-week-old rats also BP increase and coronary wall hypertrophy was found in all ontogenic periods in SHR compared to Wistar rats. Arterial wall hypertrophy was evoked by increase of CSA of medial extracellular matrix and smooth muscle cells. In 52-week-old SHR, CSA of muscle cells did not differ from that in 17-week-old SHR but the CSA of intimal and medial extracellular matrix significantly increased. The CSA of endothelial cells and CSA of intimal extracellular matrix were increased only in 52-week-old SHR. The independency between BP and trophicity of individual components of the coronary wall during ontogeny of SHR was documented.

Comparison of three volumetric techniques for estimating thyroid gland volume

The aims of this study were to estimate the preoperative thyroid volume in patients with a multinodular goiter by the use of ultrasonography (USG) and magnetic resonance imaging (MRI), and then to compare these approaches with the postsurgical total volume measured by Archimedes' principle. Materials and methods: In this study, we compared 3 methods for the determination of thyroid volume: thyroid volume measured with ellipsoid formula via 2-dimensional ultrasonography (2D USG); the stereological (point-counting) method using MRI; and the postsurgical total volume determined by the fluid displacement technique as a gold standard. Results: Thyroid volumes were calculated in a total of 20 patients (15 women and 5 men) who underwent total thyroidectomy. The mean ± SD thyroid volumes of the fluid displacement, point-counting, and ellipsoid methods were 82.75 ± 48.87, 80.45 ± 48.96, and 75.50 ± 46.59 cm^3, respectively. No significant difference was found among the methods of calculating thyroid volume (P > 0.05). The mean coefficient of error for the thyroid gland estimates derived from the technique of point-counting with MRI was under 4%. The 2D USG volume is a 10.62% underestimation of the thyroid gland volume compared with the actual volume. Conclusion: It can be concluded that there was no statistically significant difference between the 2 methods, but the 2D USG volume was underestimated; therefore, we think that the stereological method is a more efficient and reliable method than USG for thyroid gland volume estimation.

Effect of alveolex on the bone defects repair stimulated by rhBMP‐2: Histomorphometric study

The aim of this study was to evaluate histomorphometrically the effect of alveolex (Propolis 10%) on the repair of bone cavities in the calvaria of rats. A 5 mm diameter bone defect was made in the calvaria of male Wistar rats using the drill-type trephine. The defects were filled with rhBMP-2+Alveolex, rhBMP-2, Alveolex, or coagulum. Twenty-eight animals with seven subjects on each were sacrificed 30 days after surgery and samples were fixed and embedded in paraffin. Histological sections stained by HE (hematoxylin and eosin) were obtained from the calvaria bone defect and analyzed by a differential point-counting method. Group I and II, rhBMP-2+Alveolex and rhBMP-2, respectively, presented higher levels of newly formed bone than other groups (P < 0.001). There were not significant differences between groups I and II (P > 0.05). In addition, there was not significant difference between groups III and IV, Control-Coagulum and Alveolex, respectively (P > 0.05). Alveolex has increased the bone repair in calvaria defects of rats when associated to rhBMP-2, however without significant differences for rhBMP-2 isolated group; Alveolex isolated group showed the lowest levels of newly formed bone with no significant differences to coagulum group (control).

Comparison of the planimetry and point-counting methods for the assessment of the size of the mandible cysts on orthopantomograms

Objective: The purpose of this study is to compare the computer-assisted planimetry and point-counting methods in evaluating the sizes of the mandibular cysts with respect to their agreement and the time required to analyze. Study Design: The surface areas of 46 mandibular cyst lesions on orthopantomograms were estimated using the point-counting and computer-assisted planimetry methods. Three observers evaluated the outlined areas twice, using the point-counting (PC) and computer-assisted planimetry (CAP) methods with an interval of two weeks. In the planimetry technique, digitalized images and ImageJ software were used to measure the surface area of the half mandibles and cysts. The grids were superimposed over the same images and the number of points hitting the interested structures was counted for the point-counting technique. The projection area fraction (PAF) of the cysts within the mandible was estimated using the obtained values by means of the two techniques. Intraclass correlation coefficient was used to assess the level of agreement between the two methods. Inter-rater reliability analysis using the Kappa statistic was performed to determine consistency among raters. Results: CAP and PC techniques showed consistent intra-observer values in all observers. Intraclass correlation between CAP and PC measurements of first, second and third observers were found to be 0.9986, 0.9988 and 0.9994 respectively. The durations of PC technique was 32% higher than the CAP technique. Conclusion: PC and CAP methods were seemed to show sufficient agreement to be used interchangeably. The main disadvantage of the PC analysis is it takes more time than CAP method. Key words:Orthopantomogram, mandible, cyst, point-counting, planimetry.