Purkinje fibers in mammalian hearts are known to comprise the following three groups depending on their structure: group I found commonly in ungulates, group II in humans, monkeys and dogs, and group III in rodents. The aim of the present study was to document precisely the cytoarchitecture of a network of Purkinje fibers in different species by light and electron microscopy. Light microscopy of silver impregnated tissues revealed the reticular fibers ensheathing individual Purkinje strands consisting of 2-8 cells in both the ungulates (i.e., sheep and goats) and cetaceans (whales and dolphins) while they encircled each Purkinje cell in the primates (humans and monkeys), carnivores (dogs and seals), and rodents (rats). Scanning electron microscopy of NaOH digested tissues showed the ungrates (group I) to have a Purkinje fiber network composed of Purkinje strands; the cells in the strands were oval and made side-to-side and/or end-to-end connections. The Purkinje fiber network in the primates and carnivores (group II) was delicate and complicated; the Purkinje cells were usually cylindrical and connected end-to-end, the exception being their polygonal or stellate shapes at the bifurcations. Purkinje cells in the rodents (group III) resembled ventricular cardiac myocytes in cytoarchitecture. Morphologically, whales and seals respectively belonged to Purkinje cells of group I and group II. These findings indicate that the structural variety of the Purkinje fiber network may reflect the conducting function and be related to the phylogeny of the mammalian species.
{"title":"Morphological varieties of the Purkinje fiber network in mammalian hearts, as revealed by light and electron microscopy.","authors":"Noriaki Ono, Takeshi Yamaguchi, Hajime Ishikawa, Mitsue Arakawa, Naohiko Takahashi, Tetsunori Saikawa, Tatsuo Shimada","doi":"10.1679/aohc.72.139","DOIUrl":"https://doi.org/10.1679/aohc.72.139","url":null,"abstract":"<p><p>Purkinje fibers in mammalian hearts are known to comprise the following three groups depending on their structure: group I found commonly in ungulates, group II in humans, monkeys and dogs, and group III in rodents. The aim of the present study was to document precisely the cytoarchitecture of a network of Purkinje fibers in different species by light and electron microscopy. Light microscopy of silver impregnated tissues revealed the reticular fibers ensheathing individual Purkinje strands consisting of 2-8 cells in both the ungulates (i.e., sheep and goats) and cetaceans (whales and dolphins) while they encircled each Purkinje cell in the primates (humans and monkeys), carnivores (dogs and seals), and rodents (rats). Scanning electron microscopy of NaOH digested tissues showed the ungrates (group I) to have a Purkinje fiber network composed of Purkinje strands; the cells in the strands were oval and made side-to-side and/or end-to-end connections. The Purkinje fiber network in the primates and carnivores (group II) was delicate and complicated; the Purkinje cells were usually cylindrical and connected end-to-end, the exception being their polygonal or stellate shapes at the bifurcations. Purkinje cells in the rodents (group III) resembled ventricular cardiac myocytes in cytoarchitecture. Morphologically, whales and seals respectively belonged to Purkinje cells of group I and group II. These findings indicate that the structural variety of the Purkinje fiber network may reflect the conducting function and be related to the phylogeny of the mammalian species.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.139","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29026355","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
We developed a novel nano manipulator based on an atomic force microscope (AFM) that can be operated inside the sample chamber of a scanning electron microscope (SEM). This AFM manipulator is also coupled with a haptic device, and the nanometer-scale movement of the AFM cantilever can be scaled up to the millimeter-scale movement of the pen handle of the haptic device. Using this AFM manipulation system, we were able to observe the AFM cantilever and samples under the SEM and obtain topographical images of the AFM under the SEM. These AFM images contained quantitative height information of the sample that is difficult to obtain from SEM images. Our system was also useful for positioning the cantilever for accurate AFM manipulation because the manipulation scene could be directly observed in real time by SEM. Coupling of the AFM manipulator with the haptic device was also useful for manipulation in the SEM since the operator can move the AFM probe freely at any position on the sample surface while feeling the interaction force between the probe and the sample surface. We tested two types of cutting methods: simple cutting and vibration cutting. Our results showed that vibration cutting with probe oscillation is very useful for the dissection of biological samples which were dried for SEM observation. Thus, cultivated HeLa cells were successfully micro-dissected by vibration cutting, and the dissection process could be observed in real time in the SEM. This AFM manipulation system is expected to serve as a powerful tool for dissecting various biological samples at the micro and nanometer-scale under SEM observation.
{"title":"Development of a nano manipulator based on an atomic force microscope coupled with a haptic device: a novel manipulation tool for scanning electron microscopy.","authors":"Futoshi Iwata, Shinsuke Kawanishi, Hisayuki Aoyama, Tatsuo Ushiki","doi":"10.1679/aohc.72.271","DOIUrl":"https://doi.org/10.1679/aohc.72.271","url":null,"abstract":"<p><p>We developed a novel nano manipulator based on an atomic force microscope (AFM) that can be operated inside the sample chamber of a scanning electron microscope (SEM). This AFM manipulator is also coupled with a haptic device, and the nanometer-scale movement of the AFM cantilever can be scaled up to the millimeter-scale movement of the pen handle of the haptic device. Using this AFM manipulation system, we were able to observe the AFM cantilever and samples under the SEM and obtain topographical images of the AFM under the SEM. These AFM images contained quantitative height information of the sample that is difficult to obtain from SEM images. Our system was also useful for positioning the cantilever for accurate AFM manipulation because the manipulation scene could be directly observed in real time by SEM. Coupling of the AFM manipulator with the haptic device was also useful for manipulation in the SEM since the operator can move the AFM probe freely at any position on the sample surface while feeling the interaction force between the probe and the sample surface. We tested two types of cutting methods: simple cutting and vibration cutting. Our results showed that vibration cutting with probe oscillation is very useful for the dissection of biological samples which were dried for SEM observation. Thus, cultivated HeLa cells were successfully micro-dissected by vibration cutting, and the dissection process could be observed in real time in the SEM. This AFM manipulation system is expected to serve as a powerful tool for dissecting various biological samples at the micro and nanometer-scale under SEM observation.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.271","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29798172","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Waka Mitsui, Kazushi Tamura, Takeomi Mizutani, Hisashi Haga, Kazushige Kawabata
The mechanical memory effect of single cells was reported in our recent study. In order to clarify this effect, various sequential stimuli of uniaxial deformation were applied to cells by deformable culture dishes and a deformation device, and the local stiffness distribution of single C2C12 myoblasts was visualized by scanning probe microscopy. Following a single step stretching, cellular stiffness first increased steeply and then gradually decreased for two hours. By a single step stretching 30 min after a long pulse-like deformation with a pulse duration of 30 min, the cells responded in the same way. On the other hand, they did not respond to a single step stretching 30 min after a short pulse-like deformation with a pulse duration of 0.5 min. These results indicated that cellular mechanical response to external deformation is affected strongly by a preceding deformation and that the duration time of the preceding deformation is an important factor in the change in mechanical response. We consider that the change in mechanical response contributes to a regulatory mechanism of cellular contractile force.
{"title":"Mechanical response of single myoblasts to various stretching patterns visualized by scanning probe microscopy.","authors":"Waka Mitsui, Kazushi Tamura, Takeomi Mizutani, Hisashi Haga, Kazushige Kawabata","doi":"10.1679/aohc.72.227","DOIUrl":"https://doi.org/10.1679/aohc.72.227","url":null,"abstract":"<p><p>The mechanical memory effect of single cells was reported in our recent study. In order to clarify this effect, various sequential stimuli of uniaxial deformation were applied to cells by deformable culture dishes and a deformation device, and the local stiffness distribution of single C2C12 myoblasts was visualized by scanning probe microscopy. Following a single step stretching, cellular stiffness first increased steeply and then gradually decreased for two hours. By a single step stretching 30 min after a long pulse-like deformation with a pulse duration of 30 min, the cells responded in the same way. On the other hand, they did not respond to a single step stretching 30 min after a short pulse-like deformation with a pulse duration of 0.5 min. These results indicated that cellular mechanical response to external deformation is affected strongly by a preceding deformation and that the duration time of the preceding deformation is an important factor in the change in mechanical response. We consider that the change in mechanical response contributes to a regulatory mechanism of cellular contractile force.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.227","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29798167","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Anika S Mostaert, Rowena Crockett, Graham Kearn, Izhack Cherny, Ehud Gazit, Louise C Serpell, Suzanne P Jarvis
Amyloid fibrils are primarily known in a pathogenic context for their association with a wide range of debilitating human diseases. Here we show a marine invertebrate (Entobdella soleae) utilizes functional amyloid fibrils comparable to those of a unicellular prokaryote (Escherichia coli). Thioflavin-T binding and Raman spectroscopy provided evidence for the presence of amyloid in the adhesive of Entobdella soleae. We elucidated that for these two very different organisms, amyloid fibrils provide adhesive and cohesive strength to their natural adhesives. Comparing the nanoscale mechanical responses of these fibrils with those of pathogenic amyloid by atomic force microscopy revealed that the molecular level origin of the cohesive strength was associated with the generic intermolecular β-sheet structure of amyloid fibrils. Functional adhesive residues were found only in the case of the functional amyloid. Atomic force microscopy provided a useful means to characterize the internal structural forces within individual amyloid fibrils and how these relate to the mechanical performance of both functional and pathogenic amyloid. The mechanistic link of amyloid-based cohesive and adhesive strength could be widespread amongst natural adhesives, irrespective of environment, providing a new strategy for biomimicry and a new source of materials for understanding the formation and stability of amyloid fibrils more generally.
{"title":"Mechanically functional amyloid fibrils in the adhesive of a marine invertebrate as revealed by Raman spectroscopy and atomic force microscopy.","authors":"Anika S Mostaert, Rowena Crockett, Graham Kearn, Izhack Cherny, Ehud Gazit, Louise C Serpell, Suzanne P Jarvis","doi":"10.1679/aohc.72.199","DOIUrl":"https://doi.org/10.1679/aohc.72.199","url":null,"abstract":"<p><p>Amyloid fibrils are primarily known in a pathogenic context for their association with a wide range of debilitating human diseases. Here we show a marine invertebrate (Entobdella soleae) utilizes functional amyloid fibrils comparable to those of a unicellular prokaryote (Escherichia coli). Thioflavin-T binding and Raman spectroscopy provided evidence for the presence of amyloid in the adhesive of Entobdella soleae. We elucidated that for these two very different organisms, amyloid fibrils provide adhesive and cohesive strength to their natural adhesives. Comparing the nanoscale mechanical responses of these fibrils with those of pathogenic amyloid by atomic force microscopy revealed that the molecular level origin of the cohesive strength was associated with the generic intermolecular β-sheet structure of amyloid fibrils. Functional adhesive residues were found only in the case of the functional amyloid. Atomic force microscopy provided a useful means to characterize the internal structural forces within individual amyloid fibrils and how these relate to the mechanical performance of both functional and pathogenic amyloid. The mechanistic link of amyloid-based cohesive and adhesive strength could be widespread amongst natural adhesives, irrespective of environment, providing a new strategy for biomimicry and a new source of materials for understanding the formation and stability of amyloid fibrils more generally.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.199","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29798255","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
This study examined muscular atrophy and the recovery process induced by hindlimb unloading and joint immobilization in the rat soleus and plantaris muscles. Rats were divided into control, hindlimb unloading (HU), hindlimb unloading with ankle joint immobilization at the maximum dorsiflexion (HUD), and maximum plantarflexion (HUP) groups. The hindlimb was reloaded after fourteen days of unloading, and muscle atrophy and walking ability were assessed at 0, 3, and 7 days of reloading. A cross sectional area of muscle fibers in the soleus muscle on day 0 of reloading revealed sizes in order from the control, HUD, HUP down to the HU group, indicating that the HU group was the most atrophied among the four groups. These values in the plantaris muscle ranged in order from the control, HU, HUD, to HUP groups, the HUP group being the most atrophied among the four groups. These muscles recovered from atrophy in the same descending order, and the values in the HUD and HUP groups slowly recovered during the reloading periods. The HUD and HUP groups showed a central core lesion and reloading-induced lesions in some type I muscle fibers after the immobilization and reloading, one possible reason for the delayed recovery in these groups. The muscle atrophy in the HU, HUD, and HUP groups remained at day 7 although the walking ability appeared to be normal. Accordingly, further rehabilitation therapy might be necessary even if the functional ability appears to be normal.
{"title":"Influence of fixed muscle length and contractile properties on atrophy and subsequent recovery in the rat soleus and plantaris muscles.","authors":"Naoto Fujita, Takamitsu Arakawa, Takako Matsubara, Hiroshi Ando, Akinori Miki","doi":"10.1679/aohc.72.151","DOIUrl":"https://doi.org/10.1679/aohc.72.151","url":null,"abstract":"<p><p>This study examined muscular atrophy and the recovery process induced by hindlimb unloading and joint immobilization in the rat soleus and plantaris muscles. Rats were divided into control, hindlimb unloading (HU), hindlimb unloading with ankle joint immobilization at the maximum dorsiflexion (HUD), and maximum plantarflexion (HUP) groups. The hindlimb was reloaded after fourteen days of unloading, and muscle atrophy and walking ability were assessed at 0, 3, and 7 days of reloading. A cross sectional area of muscle fibers in the soleus muscle on day 0 of reloading revealed sizes in order from the control, HUD, HUP down to the HU group, indicating that the HU group was the most atrophied among the four groups. These values in the plantaris muscle ranged in order from the control, HU, HUD, to HUP groups, the HUP group being the most atrophied among the four groups. These muscles recovered from atrophy in the same descending order, and the values in the HUD and HUP groups slowly recovered during the reloading periods. The HUD and HUP groups showed a central core lesion and reloading-induced lesions in some type I muscle fibers after the immobilization and reloading, one possible reason for the delayed recovery in these groups. The muscle atrophy in the HU, HUD, and HUP groups remained at day 7 although the walking ability appeared to be normal. Accordingly, further rehabilitation therapy might be necessary even if the functional ability appears to be normal.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.151","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29028553","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Alice Pyne, Will Marks, Loren M Picco, Peter G Dunton, Arturas Ulcinas, Michele E Barbour, Siân B Jones, James Gimzewski, Mervyn J Miles
High-speed atomic force microscopy (HS AFM) in 'contact' mode was used to image at video rate the surfaces of both calcium hydroxyapatite samples, often used as artificial dental enamel in such experiments, and polished actual bovine dental enamel in both neutral and acidic aqueous environments. The image in each frame of the video of the sample was a few micrometers square, and the high-speed scan window was panned across the sample in real time to examine larger areas. Conventional AFM images of the same regions of the sample were also recorded before and after high-speed imaging. The ability of HS AFM to follow processes occurring in liquid on the timescale of a few seconds was employed to study the dissolution process of both hydroxyapatite and bovine enamel under acidic conditions. Buffered citric acid at pH values between 3.0 and 4.0 was observed to dissolve the surface layers of these samples. The movies recorded showed rapid dissolution of the bovine enamel in particular, which proceeded until the relatively small amount of acid available had been exhausted. A comparison was made with enamel samples that had been treated in fluoride solution (1 h in 300 ppm NaF, pH 7) prior to addition of the acid; the speed of dissolution for these samples was much less than that of the untreated samples. The HS AFM used an in-house designed and constructed high-speed flexure scan stage employing a push-pull piezo actuator arrangement. The HS AFM is able to follow the large changes in height (on the micrometer scale) that occur during the dissolution process.
{"title":"High-speed atomic force microscopy of dental enamel dissolution in citric acid.","authors":"Alice Pyne, Will Marks, Loren M Picco, Peter G Dunton, Arturas Ulcinas, Michele E Barbour, Siân B Jones, James Gimzewski, Mervyn J Miles","doi":"10.1679/aohc.72.209","DOIUrl":"https://doi.org/10.1679/aohc.72.209","url":null,"abstract":"<p><p>High-speed atomic force microscopy (HS AFM) in 'contact' mode was used to image at video rate the surfaces of both calcium hydroxyapatite samples, often used as artificial dental enamel in such experiments, and polished actual bovine dental enamel in both neutral and acidic aqueous environments. The image in each frame of the video of the sample was a few micrometers square, and the high-speed scan window was panned across the sample in real time to examine larger areas. Conventional AFM images of the same regions of the sample were also recorded before and after high-speed imaging. The ability of HS AFM to follow processes occurring in liquid on the timescale of a few seconds was employed to study the dissolution process of both hydroxyapatite and bovine enamel under acidic conditions. Buffered citric acid at pH values between 3.0 and 4.0 was observed to dissolve the surface layers of these samples. The movies recorded showed rapid dissolution of the bovine enamel in particular, which proceeded until the relatively small amount of acid available had been exhausted. A comparison was made with enamel samples that had been treated in fluoride solution (1 h in 300 ppm NaF, pH 7) prior to addition of the acid; the speed of dissolution for these samples was much less than that of the untreated samples. The HS AFM used an in-house designed and constructed high-speed flexure scan stage employing a push-pull piezo actuator arrangement. The HS AFM is able to follow the large changes in height (on the micrometer scale) that occur during the dissolution process.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.209","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29798256","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Scanning probe microscopy (SPM) provides a range of strategies for studying biological phenomena due to its ability to image surfaces under liquids. However, some cellular events, such as cell migration, exceed the maximum measurable range of SPM. Recently, we have developed a wide range scanning probe microscope (WR-SPM) to investigate cellular events which exceed the range of the conventional SPM. In this review, we introduce the instrumentation of the WR-SPM, which can measure a sample for 400 µm in the xy directions and 23 µm in the z direction. We then show the application of the WR-SPM to studies of the stiffness response of epithelial cells to an external loading force and demonstrat that the stiffness of the epithelial cells increases under stretched conditions. We also showed the results on the mesh structure on the surface of a melanoma cell as well as the regulatory mechanism of the cellular contractile force by the combined use of topographical and mechanical modes of the WR-SPM. These findings indicate that the WR-SPM is very useful for studying the functions of a cell in relation to the surface structure and mechanical properties of that cell.
{"title":"Wide range scanning probe microscopy for probing mechanical effects on cellular function.","authors":"Takeomi Mizutani, Hisashi Haga, Kosaku Kato, Kazushige Kawabata","doi":"10.1679/aohc.72.235","DOIUrl":"https://doi.org/10.1679/aohc.72.235","url":null,"abstract":"<p><p>Scanning probe microscopy (SPM) provides a range of strategies for studying biological phenomena due to its ability to image surfaces under liquids. However, some cellular events, such as cell migration, exceed the maximum measurable range of SPM. Recently, we have developed a wide range scanning probe microscope (WR-SPM) to investigate cellular events which exceed the range of the conventional SPM. In this review, we introduce the instrumentation of the WR-SPM, which can measure a sample for 400 µm in the xy directions and 23 µm in the z direction. We then show the application of the WR-SPM to studies of the stiffness response of epithelial cells to an external loading force and demonstrat that the stiffness of the epithelial cells increases under stretched conditions. We also showed the results on the mesh structure on the surface of a melanoma cell as well as the regulatory mechanism of the cellular contractile force by the combined use of topographical and mechanical modes of the WR-SPM. These findings indicate that the WR-SPM is very useful for studying the functions of a cell in relation to the surface structure and mechanical properties of that cell.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.235","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29798168","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The relationship between the higher-order structure of human metaphase chromosomes and the distribution of topoisomerase IIα was analyzed by a comparison of atomic force microscope (AFM) and fluorescence microscope images of the same chromosome. AFM imaging of chromosomes in liquid revealed the presence of alternating ridges and grooves on the surfaces of the sister chromatids. In contrast, the fluorescence image of the chromosomes stained with the anti-topoisomerase IIα antibody showed that the fluorescence intensity of topoisomerase IIα was not uniform and that there were alternating strong and weak spots along the chromosome axes. A comparison of the AFM image with a fluorescence microscope image of the same chromosome further demonstrated that ridges and grooves corresponded to strong and weak fluorescence intensities of topoisomerase IIα, respectively. These findings suggest that the distribution of topoisomerase IIα has a close connection with the higher-order structure of human metaphase chromosomes.
{"title":"Structural analysis of human chromosomes by atomic force and light microscopy in relation to the distribution of topoisomerase IIα.","authors":"Osamu Hoshi, Tatsuo Ushiki","doi":"10.1679/aohc.72.245","DOIUrl":"https://doi.org/10.1679/aohc.72.245","url":null,"abstract":"<p><p>The relationship between the higher-order structure of human metaphase chromosomes and the distribution of topoisomerase IIα was analyzed by a comparison of atomic force microscope (AFM) and fluorescence microscope images of the same chromosome. AFM imaging of chromosomes in liquid revealed the presence of alternating ridges and grooves on the surfaces of the sister chromatids. In contrast, the fluorescence image of the chromosomes stained with the anti-topoisomerase IIα antibody showed that the fluorescence intensity of topoisomerase IIα was not uniform and that there were alternating strong and weak spots along the chromosome axes. A comparison of the AFM image with a fluorescence microscope image of the same chromosome further demonstrated that ridges and grooves corresponded to strong and weak fluorescence intensities of topoisomerase IIα, respectively. These findings suggest that the distribution of topoisomerase IIα has a close connection with the higher-order structure of human metaphase chromosomes.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.245","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29798169","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
A disintegrin and metalloproteinase with thrombospondin motifs 9 (ADAMTS9) is known to influence aggrecan degradation in endochondral ossification, but its role has not been well understood. In the present study, in vitro gene expression of ADAMTS9 was investigated by RT-PCR in ATDC5 cells in which experimentally chondrogenic differentiation had been induced. We also investigated the protein localization and gene expression pattern of ADAMTS9 in the tibia growth plate cartilage of male mice in a day 1 neonate, 7-week-old young adult, and a 12-week-old adult by immunohistochemistry and in situ hybridization and compared the results with the expression of proliferating cell nuclear antigen (PCNA) and type X collagen for the identification of proliferative and hypertrophic chondrocyte phenotypes, respectively. We found the gene expression of ADAMTS9 by ATDC5 cells as a dual mode, both before the expression of type X collagen and after hypertrophic differentiation. The immunoreactivity of ADAMTS9 was observed in chondrocytes of proliferative and hypertrophic zones in the growth plate. The population of ADAMTS9 positive cells decreased with age. The results of the present study suggest that ADAMTS9 might have a role in aggrecan cleavage around the chondrocytes to allow chondrocyte proliferation and hypertrophy.
{"title":"A disintegrin and metalloproteinase with thrombospondin motifs 9 (ADAMTS9) expression by chondrocytes during endochondral ossification.","authors":"Kanae Kumagishi, Keiichiro Nishida, Tomoichiro Yamaai, Ryusuke Momota, Shigeru Miyaki, Satoshi Hirohata, Ichiro Naito, Hiroshi Asahara, Yoshifumi Ninomiya, Aiji Ohtsuka","doi":"10.1679/aohc.72.175","DOIUrl":"https://doi.org/10.1679/aohc.72.175","url":null,"abstract":"<p><p>A disintegrin and metalloproteinase with thrombospondin motifs 9 (ADAMTS9) is known to influence aggrecan degradation in endochondral ossification, but its role has not been well understood. In the present study, in vitro gene expression of ADAMTS9 was investigated by RT-PCR in ATDC5 cells in which experimentally chondrogenic differentiation had been induced. We also investigated the protein localization and gene expression pattern of ADAMTS9 in the tibia growth plate cartilage of male mice in a day 1 neonate, 7-week-old young adult, and a 12-week-old adult by immunohistochemistry and in situ hybridization and compared the results with the expression of proliferating cell nuclear antigen (PCNA) and type X collagen for the identification of proliferative and hypertrophic chondrocyte phenotypes, respectively. We found the gene expression of ADAMTS9 by ATDC5 cells as a dual mode, both before the expression of type X collagen and after hypertrophic differentiation. The immunoreactivity of ADAMTS9 was observed in chondrocytes of proliferative and hypertrophic zones in the growth plate. The population of ADAMTS9 positive cells decreased with age. The results of the present study suggest that ADAMTS9 might have a role in aggrecan cleavage around the chondrocytes to allow chondrocyte proliferation and hypertrophy.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.175","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29028555","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Raphael Imer, Terunobu Akiyama, Nico F de Rooij, Martin Stolz, Ueli Aebi, Niklaus F Friederich, Urs Staufer
We have recently demonstrated that indentation-type atomic force microscopy (IT-AFM) is capable of detecting early onset osteoarthritis (OA) (Stolz, 2009). This study was based on biopsies, using a desk-top commercial atomic force microscope (AFM). However, cartilage analysis in the knee joints needs to be non-destructive to avoid new seeding points for OA by the taking of biopsies. This requires bringing the probe tip in contact with the articular cartilage (AC) surface inside the joint. Here we present our recent progress towards a medical instrument for performing such IT-AFM measurements for in-vivo knee diagnostics. The scanning force arthroscope (SFA) integrates a miniaturized AFM into a standard arthroscopic sleeve, and is used for direct, quantitative, in situ inspection of AC (Imer et al., 2006). The stabilization and the positioning of the instrument relative to the surface under investigation were performed by means of eight inflatable balloons. An integrated three-dimensional, piezoelectric scanner allowed raster scanning and probing of a small area of cartilage around the point of insertion. An AFM probe with an integrated deflection sensor was mounted at the distal end of the instrument. Using this instrument, several measurements were performed on agarose gel and on porcine cartilage samples. The load-displacement curves obtained were analyzed and the dynamic elastic moduli | E(*) | were calculated. A good correlation between these values and those published in the scientific literature was found. Therefore, we concluded that the SFA can provide quantitative measurements to detect early pathological changes in OA.
我们最近证明,压痕型原子力显微镜(IT-AFM)能够检测早发性骨关节炎(OA) (Stolz, 2009)。这项研究是基于活组织检查,使用台式商用原子力显微镜(AFM)。然而,膝关节的软骨分析需要是非破坏性的,以避免活检产生新的骨关节炎播散点。这需要将探针尖端与关节内的关节软骨(AC)表面接触。在这里,我们介绍了我们最近的进展,医疗仪器执行这样的IT-AFM测量在体内膝关节诊断。扫描力关节镜(SFA)将小型AFM集成到标准关节镜套筒中,用于直接、定量、原位检查AC (Imer et al., 2006)。仪器相对于被测地表的稳定和定位是通过8个充气气球来实现的。集成的三维压电扫描仪可以对插入点周围的一小块软骨进行光栅扫描和探测。在仪器的远端安装了一个集成偏转传感器的AFM探针。利用该仪器,对琼脂糖凝胶和猪软骨样品进行了多次测量。对得到的载荷-位移曲线进行了分析,并计算了动弹性模量| E(*) |。研究发现,这些数值与科学文献中发表的数值之间存在良好的相关性。因此,我们认为SFA可以为检测OA的早期病理变化提供定量测量。
{"title":"The measurement of biomechanical properties of porcine articular cartilage using atomic force microscopy.","authors":"Raphael Imer, Terunobu Akiyama, Nico F de Rooij, Martin Stolz, Ueli Aebi, Niklaus F Friederich, Urs Staufer","doi":"10.1679/aohc.72.251","DOIUrl":"https://doi.org/10.1679/aohc.72.251","url":null,"abstract":"<p><p>We have recently demonstrated that indentation-type atomic force microscopy (IT-AFM) is capable of detecting early onset osteoarthritis (OA) (Stolz, 2009). This study was based on biopsies, using a desk-top commercial atomic force microscope (AFM). However, cartilage analysis in the knee joints needs to be non-destructive to avoid new seeding points for OA by the taking of biopsies. This requires bringing the probe tip in contact with the articular cartilage (AC) surface inside the joint. Here we present our recent progress towards a medical instrument for performing such IT-AFM measurements for in-vivo knee diagnostics. The scanning force arthroscope (SFA) integrates a miniaturized AFM into a standard arthroscopic sleeve, and is used for direct, quantitative, in situ inspection of AC (Imer et al., 2006). The stabilization and the positioning of the instrument relative to the surface under investigation were performed by means of eight inflatable balloons. An integrated three-dimensional, piezoelectric scanner allowed raster scanning and probing of a small area of cartilage around the point of insertion. An AFM probe with an integrated deflection sensor was mounted at the distal end of the instrument. Using this instrument, several measurements were performed on agarose gel and on porcine cartilage samples. The load-displacement curves obtained were analyzed and the dynamic elastic moduli | E(*) | were calculated. A good correlation between these values and those published in the scientific literature was found. Therefore, we concluded that the SFA can provide quantitative measurements to detect early pathological changes in OA.</p>","PeriodicalId":8307,"journal":{"name":"Archives of histology and cytology","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2009-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1679/aohc.72.251","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"29798170","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}