Method of forming an image of cilium and cilia

Details Method of forming an image of cilium and cilia Method of forming an image of cilium and cilia

2001-04-25

2004-09-14

Johns, Andrew W. (Department: 2621)

Image analysis

Applications

Biomedical applications

C600S160000

Reexamination Certificate

active

06792138

ABSTRACT:

This application claims benefit of Japanese Application No. 2000-128968 filed on Apr. 25, 2000, the contents of which are incorporated by this reference.
FIELD OF THE INVENTION
This invention relates to a method of forming a detailed image of cilium and cilia which exists in an epithelial layer of a cellular structure of a bronchi or airway.
BACKGROUND OF THE INVENTION
As for mucociliary transport, ciliated epithelial cells play an important role as a barrier system for airway structures, such as bronchi or nasal cavities. The ciliated epithelial cells cover the surface of the bronchi. Thus, the abnormalities of ciliated epithelial cells cause various disorders of the trachea bronchial tree.
Ciliated epithelial cells are explained with reference to
FIGS. 5 and 6
. The cross-sectional view of a layer structure of a bronchial wall is shown in FIG.
5
. Moreover, a partial cross-sectional view of a ciliated epithelial cells tissue is shown in FIG.
6
.
As shown in
FIG. 5
, the bronchial wall consists of an epithelial layer
33
which has ciliated columnar epithelial cells
31
and a basement membrane
32
, a subepithelial layer
35
which has bundles of elastic fibers
34
, a muscle layer
37
which has a smooth muscle (bundle)
36
, and an outer muscle layer
39
which is the outer layer of the bronchial wall. The cartilaginous layer
41
consists of cartilage
40
, a pericartilaginous layer
42
and bronchial glands
38
.
As shown in
FIG. 6
, the epithelial layer
33
consists of a basement membrane
32
, ciliated columnar epithelial cells
31
and cilium
43
, with the top of the epithelial layer
33
covered with the gel-like mucus
44
. Ciliated columnar epithelial cells
31
are transparent cylinder shaped cells having a nucleus
45
and a length of about 100 &mgr;m. The ciliated columnar epithelial cells
32
exist on the basement membrane
32
.
Cilium
43
is a colorless and transparent hair-like object with a diameter of 0.1-0.2 &mgr;m and length of several &mgr;m. Several hundred strands exist at the upper part of each ciliated columnar epithelial cell
31
. The gel-like mucus
44
exists from the upper part of the ciliated columnar epithelial cells
31
extended upward by about 6 &mgr;m. Furthermore, the upper part of mucus
44
is solid-like. Cilium
43
performs the ciliary motion, a repeated transverse oscillation in the shape of a whip, in the gel-like mucus
44
.
Since the epithelial layer
33
is transparent, when the bronchial epithelium of mucus in an organism is observed using an endoscope optical system, an illumination light permeates the cellular structure containing the epithelial layer
33
by about 0.5 mm. For this reason, the bundles of elastic fibers
34
which have color and exist in the cellular structure of a sublayer outside of the epithelial layer
33
, are actually observed.
Apart from that, cilium disappears when a cell of bronchial epitheliums becomes cancerous. For this reason, the disappearance of cilium plays an important role in the evaluation of a malignant cell, and location of an affected region.
However, as mentioned above, since ciliated epithelial cells are transparent, it is hard in a conventional method to observe a disappearance of cilium in the process of the cells becoming cancerous, with the naked eye.
The following conventional methods have been used to observe cilia. There is the fluorescent method for observing with an endoscope, wherein an cellular organization extracted from an living body is dyed with a fluorescent paint and a laser light is irradiated onto the cellar organization. There is the method of observing a motion state more indirectly according to a change in the transmittance of a light and the measured frequency of the cilium. There is also the method of observing the scattered light of cilium by the transmitted illumination using a culture cell. In addition, there is the method of observing a change in the input-output signals using the light.
Currently, although it turns out that cilium disappears in the process of becoming cancer, changes in the form and the motion of the cilium caused by other diseases have not been clarified. This is because observation of cilium of a living tissue by the endoscope, etc. was difficult. If a change in the form or motion of cilium of and cilia a living tissue can be clearly discerned and data can be stored, such will become a great assistance in identifying or diagnosing diseases. This is not limited to the specialty of a bronchi or trachea bronchial tree but can also be applied to an otorhinilogy specialty and an obstetrics and gynecology specialty as well.
Moreover, although the observation of a change of the ciliary motion has only been performed in a culture cell, if a cilium observation by the naked eye of an in vivo becomes possible, it could not only confirm the effect of a medicine, but it could also contribute greatly to new medicine development in animal experiments.
Thus, when studying the relationship with a disease, it is important that the ciliary motion in the living body be observed as a direct image.
Conventionally, observing cilium in the living body with the naked eye has not been thought of at all, as mentioned above. In the conventional methods, a microscope, etc. was used to observe a partial cellular structure only after it was collected from an organism and moved to a receptacle so that the cilium of a cellular structure could be observed with the naked eye. However, according to the conventional methods, since the object was observed only about extremely limited parts, the exact range of a wide range observation and a disease could not be specified.
Moreover, since it is indispensable to collect the cellular structure which then becomes an observation object, the physical burden on a subject and the observer's operation burden becomes significant.
In view of the above mentioned problems inherent in the prior art, it is an object of the present invention to provide a method of forming an image of cilium and its motion in vivo, which can be observed with the naked eye.
SUMMARY OF THE INVENTION
Preferred embodiments of the present invention will now be described.
As mentioned above, in an endoscopic observation using any of the conventional observation procedures, the motion of cilium in vivo has not been confirmed with the naked eye.
In an observation screen, a white luminescent point like a halation may be visible accidentally. When it was observed that by raising the observation magnifying power stepwise about this white luminescent point, it has been confirmed that the white luminescent point part was shaking on and off at more than a predetermined magnifying power.
In the conventional observation, an observer had no idea that he was observing a luminescent point at all, since a white luminescent point is recognized as an obstacle to an observation of a subepithelial layer part, and the observation magnifying power in the endoscope was not high enough to confirm the shake of the luminescent point.
However, when this white luminescent point was observed by raising the magnifying power, it was confirmed that this white shake of a luminescent point was none other than the cilium and cilia with motions of the mucous membrane layer due to the motion of the cilia.
The confirmation of the motion of the cilium in vivo by the an endoscope was a very significant achievement scientifically because this was the first time ever that this has been accomplished.
Moreover, an animal and a person's extracted bronchi were observed by using a stereoscopic microscope. When a stereoscopic microscope was used, it was found that a higher magnifying power (500× on a 14 inch monitor) than that of an endoscope was needed in order to observe the cilium. Specifically, the structure (cilium) which by moving causes a halation has been confirmed by further enlarging the part which causes the halation during an animal experiment. In addition, the characteristics of periodicity and polarity were observed in the motion of this structure.
Therefore, cilium can be

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