BackgroundGrowing demand for three dimensional (3D) digital images of embryos for purposes of phenotypic assessment drives implementation of new histological and imaging techniques. Among these micro-computed tomography (μCT) has recently been utilized as an effective and practical method for generating images at resolutions permitting 3D quantitative analysis of gross morphological attributes of developing tissues and organs in embryonic mice. However, histological processing in preparation for μCT scanning induces changes in organ size and shape. Establishing normative expectations for experimentally induced changes in size and shape will be an important feature of 3D μCT-based phenotypic assessments, especially if quantifying differences in the values of those parameters between comparison sets of developing embryos is a primary aim. Toward that end, we assessed the nature and degree of morphological artifacts attending μCT scanning following use of common fixatives, using a two dimensional (2D) landmark geometric morphometric approach to track the accumulation of distortions affecting the embryonic head from the native, uterine state through to fixation and subsequent scanning.ResultsBouin's fixation reduced average centroid sizes of embryonic mouse crania by approximately 30% and substantially altered the morphometric shape, as measured by the shift in Procrustes distance, from the unfixed state, after the data were normalized for naturally occurring shape variation. Subsequent μCT scanning produced negligible changes in size but did appear to reduce or even reverse fixation-induced random shape changes. Mixtures of paraformaldehyde + glutaraldehyde reduced average centroid sizes by 2-3%. Changes in craniofacial shape progressively increased post-fixation.ConclusionsThe degree to which artifacts are introduced in the generation of random craniofacial shape variation relates to the degree of specimen dehydration during the initial fixation. Fixation methods that better maintain original craniofacial dimensions at reduced levels of dehydration and tissue shrinkage lead to the progressive accumulation of random shape variation during handling and data acquisition. In general, to the degree that embryonic organ size and shape factor into μCT-based phenotypic assessments, procedurally induced artifacts associated with fixation and scanning will influence results. Experimental designs will need to address these significant effects, either by employing alternative methods that minimize artifacts in the region of focus or in the interpretation of statistical patterns.

Optical coherence tomography (OCT) imaging has become a standard diagnostic tool in ophthalmology, providing essential information associated with various eye diseases. In order to investigate the dynamics of the ocular fundus, we present a simple and accurate automated algorithm to segment the inner limiting membrane in video-rate optic nerve head spectral domain (SD) OCT images. The method is based on morphological operations including a two-step contrast enhancement technique, proving to be very robust when dealing with low signal-to-noise ratio images and pathological eyes. An analysis algorithm was also developed to measure neuroretinal tissue deformation from the segmented retinal profiles. The performance of the algorithm is demonstrated, and deformation results are presented for healthy and glaucomatous eyes.

A 39-year-old woman presented to the emergency department 3 times over 6 weeks with abdominal pain, constipation, nausea and vomiting. She had a medical history of hypothyroidism and infertility, and was taking levothyroxine, follitropin alfa injections and folic acid. She lived in a condominium and worked in an office. She did not drink alcohol and was a nonsmoker. She did not have a notable family history.On her third visit to the emergency department, the patient reported abdominal pain, fatigue, nausea and vomiting. Her vital signs were normal. Her hemoglobin level was 67 (normal 115-155) g/L with a mean corpuscular volume of 88.5 (normal 80.0-98.0) fL. She was admitted to hospital to investigate the anemia and possible gastrointestinal bleeding. Her blood film showed basophilic stippling, slight microcytosis and hypochromasia with no hemolysis. Levels of electrolytes, calcium, magnesium, phosphate, vitamin B 12 , thyroid-stimulating hormone and liver enzymes were normal. Cultures of blood and urine samples were negative. Results of esophagogastroduodenoscopy, colonoscopy and computed tomography and ultrasonography of her abdomen and pelvis were normal. A bone marrow biopsy to investigate the anemia showed a moderate to marked increase in iron storage. Diagnostic laparoscopy for possible endometriosis showed no visible endometriosis. Biopsies of simple ovarian cysts were sent for pathologic examination. The patient's abdominal pain improved and her hemoglobin remained higher than 70 g/L after transfusions. Her discharge diagnosis was anemia of unknown cause and possible mild endometriosis. She was prescribed analgesics, laxatives and her usual medications, and was scheduled for follow-up with the internal medicine and hematology services.At the outpatient follow-up with the internal medicine service 2 weeks after discharge, the patient reported improved abdominal pain but ongoing fatigue, shortness of breath, headaches and tinnitus. A urine sample was sent out for porphyria screening. At the second follow-up 4 weeks after discharge, at which point one of the authors became involved (H.A.), her porphyria screen showed elevated levels of coproporphyrin III and δ-aminolevulinic acid. Her biopsies did not show endometriosis. On further inquiry, the patient reported having taken Ayurvedic medications daily to treat infertility for more than a year. Her regimen varied, ranging from a few to a dozen pills daily. She had stopped taking them before her admission to hospital because of the abdominal pain, but had resumed them after discharge. Her blood lead level was 55 (normal < 2) µg/dL. A history of the patient's occupational and environmental exposure did not reveal other potential sources of lead exposure.