Perturbation of mammalian cell division. III. The topography and kinetics of extrusion subdivision.
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If mitotic-arrested, cold-stored HeLa cells are incubated at 37 degrees C a proportion of the population divides by an aberrant process which we have called subdivision by extrusion. This process has been studied by time-lapse photography and shown to differ from normal cleavage in several respects. The cell surface becomes more generally mobile and, instead of producing the precisely localized furrowing activity of cytokinesis, gives rise to multiple surface protrusions. These protrusions enlarge at the expense of the parent cell and develop into a cluster of small daughter cells (mini segregants). The surface structure of the cell, as seen by scanning electron microscopy, also changes; the microvilli characteristic of interphase, metaphase and cleaving HeLa cells are lost during extrusion and the cell surface becomes smooth. Extrusion activity is much more variable than division by cleavage in terms of both topography and kinetics, and in general takes longer to complete. Some cells in the cold-treated populations divide by mixtures of cleavage and extrusion or by cleavage alone. The relative numbers of cells dividing in different ways vary with the conditions of pretreatment and incubation of the mitotic cells. The greater the perturbation (e.g. longer cold storage), the greater the proportion of extruding rather than cleaving cells. Human diploid cells can also be induced to subdivide by extrusion. Possible mechanisms underlying the different types of division activity are discussed. (+info)
A vital stain for studying membrane dynamics in bacteria: a novel mechanism controlling septation during Bacillus subtilis sporulation.
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At the onset of sporulation in Bacillus subtilis, two potential division sites are assembled at each pole, one of which will be used to synthesize the asymmetrically positioned sporulation septum. Using the vital stain FM 4-64 to label the plasma membrane of living cells, we examined the fate of these potential division sites in wild-type cells and found that, immediately after the formation of the sporulation septum, a partial septum was frequently synthesized within the mother cell at the second potential division site. Using time-lapse deconvolution microscopy, we were able to watch these partial septa first appear and then disappear during sporulation. Septal dissolution was dependent on sigma E activity and was partially inhibited in mutants lacking the sigma E-controlled proteins SpoIID, SpoIIM and SpoIIP, which may play a role in mediating the degradation of septal peptidoglycan. Our results support a model in which sigma E inhibits division at the second potential division site by two distinct mechanisms: inhibition of septal biogenesis and the degradation of partial septa formed before sigma E activation. (+info)
Iron-induced cytotoxicity in cultured rat retinal neurons.
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Oxidative stress has been proposed as a major injury mechanism in the central nervous system including the retina. In this study, as an initial attempt to study the mechanism of oxidative injury in the retina, we developed a cell culture model by utilizing the iron exposure paradigm. Exposure of rat retinal cultures for 24 hours to 10-40 MicroM ferrous or ferric chloride induced a concentration-dependent death of retinal neurons but not of photoreceptors or astrocytes. An antioxidant, trolox effectively attenuated the iron-induced death of neurons and photoreceptors in a dose-dependent manner whereas neither glutamate receptor antagonists nor cycloheximide were protective. Of retinal interneurons, GABAergic neurons were more vulnerable to the iron toxicity than calbindin (+) horizontal neurons. These findings show that iron exposure induces anti-oxidant-sensitive neuronal injury in retinal culture, independent of the excitotoxic or the apoptotic mechanisms. Of retinal neurons, different cell types exhibit differential vulnerabilities to the iron-induced oxidative injury. This simplified culture model system may be useful in elucidating mechanisms of oxidative injury in the retina. (+info)
Quantitative analysis of transforming growth factor beta 1 and 2 in ovarian carcinoma.
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Transforming growth factor beta (TGF-beta) is an important family of cytokines that may promote tumor growth in vivo through several mechanisms including interference with antitumor T-cell immune responses, alteration of factors in the stroma and matrix, and the promotion of angiogenesis. TGF-beta isotypes have been detected in malignant and normal ovarian tissues. We have determined by quantitative immunohistochemistry the density of TGF-beta1, TGF-beta2, and human leukocyte antigen (HLA) Class I and Class II antigens on malignant cells in paired primary and metastatic specimens from 10 patients with ovarian carcinoma. Cryostat sections of specimens from the carcinomas and from normal ovaries of three women of similar age without ovarian cancer were stained respectively with specific antibodies to TGF-beta1, TGF-beta2, and HLA Class I and II antigens, and with isotype-matched control antibodies. Antigen density was quantitated blindly as mean absorbance on a SAMBA 4000 image analyzer. TGF-beta1 and TGF-beta2 were overexpressed in both primary and metastatic tumor specimens in comparison with normal ovarian tissue. No statistical correlation was found between the expression of TGF-beta1 or TGF-beta2 and HLA class I or HLA class II, which suggests that TGF-beta isotypes could have effects on the immune system other than down-modulation of these HLA molecules. Furthermore, the lack of association between levels of TGF-beta expression and the reduced expression of HLA molecules could suggest that tumor cells expressing both HLA and TGF-beta may be suitable targets for adaptive immunotherapy. Additional studies are necessary to determine whether TGF-beta expressed by ovarian cancer cells merits evaluation as a therapeutic target. (+info)
Effect of cerebrospinal fluid shunting on experimental syringomyelia: magnetic resonance imaging and histological findings.
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The histological changes associated with syringomyelia after reduction of the syrinx size were investigated after cerebrospinal fluid shunting in experimental syringomyelia in the rabbit. Five weeks after syringomyelia was induced by the injection of kaolin into the cisterna magna in Japanese white rabbits, ventriculosubgaleal shunting or syringoepidural shunting were performed. After 1 week magnetic resonance (MR) imaging and histological examination were then carried out. Five of 11 shunted animals showed postoperative reduction of syrinx size on MR imaging. Grossly, some specimens showed cavity collapse and parenchymal healing, and others showed a small residual syrinx in the dorsal horn. The most dramatic histological changes occurred in the gray matter. Specimens with syrinx collapse showed rarefaction and tearing of the gray matter, with mild glial reaction. The edematous gray matter showed both degeneration and regeneration, with neuronal processes surrounded by edema fluid. Reactive astrocytes were observed mainly at the margin of the residual syrinx. Some astrocytic processes invested the extraaxonal space and gray matter lacked supportive tissue. Greater reduction of the syrinx after shunting operation was correlated with more regeneration and less degeneration, and the white matter was edematous and histological changes were milder. Syrinx shrinkage occurred after shunting in this experimental model of syringomyelia. The selective vulnerability of gray matter even after shunting may explain discrepancies between imaging findings and clinical features in this disease. The study supports the potential benefit from early treatment, considering the associated morphological findings of regeneration. (+info)
Mitral annular dilatation and papillary muscle dislocation without mitral regurgitation in sheep.
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BACKGROUND: Asymmetrical mitral annular (MA) dilatation and papillary muscle dislocation are implicated in the pathogenesis of functional mitral regurgitation (MR). METHODS AND RESULTS: To determine the mechanism by which annular and papillary muscle geometric alterations result in MR, we implanted radiopaque markers in the left ventricle, mitral annulus, anterior and posterior mitral leaflets, and papillary muscle tips and bases in 2 groups of sheep. One group served as controls (CTL, n=7); an experimental group (EXP, n=9) underwent topical phenol application to obliterate anterior annular and leaflet muscle (confirmed histologically ex vivo). After 1 week of recovery, markers were imaged with biplane videofluoroscopy, and hemodynamic data were recorded. MA area (computed from 3-dimensional marker coordinates) was 11% to 13% larger in the EXP group than in the CTL group (P<0.05 by ANOVA). This area increase resulted exclusively from intercommissural axis increase except in 1 heart with large (>1 cm) increases in both the intercommissural and septolateral annular axes. The anterior papillary muscle tip in EXP was displaced from CTL by 2.9+/-0.23 mm toward the anterolateral left ventricle and 2.5+/-0.12 mm toward the mitral annulus at end systole; the posterior papillary muscle geometry was unchanged. Transthoracic echocardiography revealed MR only in the heart exhibiting biaxial annular enlargement. CONCLUSIONS: MA dilatation in the intercommissural dimension with anterior papillary muscle tip displacement toward the annulus is insufficient to produce MR in sheep. Functional MR may require MA dilatation in the septolateral axis, as observed with proximal circumflex coronary occlusion. (+info)
Subchondral insufficiency fracture of the femoral head: a differential diagnosis in acute onset of coxarthrosis in the elderly.
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OBJECTIVE: To document subchondral insufficiency fracture (SIF) of the femoral head and investigate its frequency. METHODS: The study was based on a retrospective review of 464 removed femoral heads (from 419 patients) with both radiologic and histologic evidence of subchondral collapse. Gross photographs, specimen radiographs, and histologic sections were reevaluated in all cases. Available clinical notes and imaging studies were also reviewed. RESULTS: Ten cases previously diagnosed as osteonecrosis were reinterpreted as SIF on a histopathologic basis. All of these patients were women over 65 years old (average age 75) with osteopenia. The initial symptom was acute onset of hip pain. Radiologically, a subchondral collapse, mainly in the superolateral segment of the femoral head, was noted. Magnetic resonance imaging, available in 3 cases, showed diffuse low intensity on T1-weighted images and high intensity on T2-weighted or fat-suppressed images. Bone scintigraphy, available in 4 cases, showed increased uptake in the femoral head. Histopathologically, a 1.0-2.5-cm long linear whitish gray zone, comprising fracture callus and granulation tissue, was found beneath the subchondral bone end plate. There was no evidence of antecedent osteonecrosis. CONCLUSION: The results of this study indicate that SIF should be included in the differential diagnosis of acute onset of coxarthrosis in the elderly. (+info)
Correlation of bone scintigraphy and histological findings in medial tibial syndrome.
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OBJECTIVE: To correlate bone scintigraphy and histopathological findings in patients with medial tibial syndrome. METHODS: Twenty patients (32 limbs) with a clinical diagnosis of medial tibial syndrome had surgery. Bone scintigraphy before the operation was compared with the histological appearance of bone and periosteal specimens obtained at surgery. RESULTS: Delayed bone scintigraphy showed normal appearance in 11 limbs, characteristic diffuse tubular pattern uptake in 16 limbs, and focal uptake in five. Periosteal histology disclosed fibrous thickening as the most common finding associated with increased vascularity, occasionally with chronic inflammatory cell infiltration, haemosiderin, and acid mucopolysaccharide deposition. Loss of osteocytes was the main finding of bone histology associated with some enlargement of lacunae and lamellar structure disruption. A grading system was used to score normal and abnormal histological appearance. For analysis the findings were regrouped to provide tables using Fisher's exact test. There was no correlation between bone scintigraphy and the histology of bone and periosteum, but two interesting observations were noted. Those cases with periosteal thickening had mostly normal bone scan appearance (p = 0.0028). Those cases with low levels of osteocyte loss had mostly abnormal bone scintigraphy. CONCLUSION: Abnormal histological appearance of bone and periosteum is a feature of medial tibial syndrome. These histological findings show poor correlation with bone scintigraphy. The exact pathogenesis of this syndrome remains unclear. (+info)