As for the mechanism of formation of connective tissue fibres, much had been left to be further studied and solved. To clarify this problem, the following close observation of the development of the primordium of the cutis of amphibians was conducted: this investigation being finally proved to be most effective and proper for throwing clear light upon this subject. The process of the investigation undertaken is as follows:
The ectoderm of younger larvae of
Rana Japonica, after being immersed in the dilute solution of chloretone for about ten minutes, can be stripped off the body. Then appears a thin semitransparent membranous substance which covers the mesoderm. This is the primordium of the cutis.
After fixation in formaline, the membranous substance is stripped off the body surface as a film-sample. Next this is stained with iron-hematoxyline or, mounted with solution of dry human plasma without being stained at all, is observed under a phase-contrast-microscope (manufactured by Tiyoda company, Bright contrast, High).
In the early stage the thin membranous substance is filled with irregularly arranged fine granules (Fig. 2). In the stage 3 (Fig. 1) the fine granules are arranged in one definite direction, parallel to the body surface (Fig. 3). In the next stage (stage 3-5, Fig. 1) they are rearranged in two directions intersecting each other, forming a latticework (Fig. 4, 5). The author names this condition of the granules “prefibrils.”
The characteristic patterns such as triangles and pentagons which are typically seen in the cutis of amphibians and other animals are already formed in this stage (Fig. 7).
The “prefibrils” are gradually developed into genuine fibrils in the inner-gill-stage (7, Fig. 1). But they still remain invariably unstainable even though the methods of dyeing collagenous fibers employed by VAN GIESON, MALLORY and BIELSCHOWSKY are adopted, and again they show no double refractions. They are digested by neither pepsin nor trypsin. In the stage 8 (Fig. 1) the fibrils acquire both affinity for staining the collagenous fibre and slight optical polarity, but they still hold resistance to the digestive process of pepsin and trypsin. In the later stage of metamorphosis, the cutis fibres come to acquire for the first time the characteristic nature of a genuine collagen. Therefore, the cutis fiberes before the stage of metamorphosis can be defined to belong to the stage of “precollagenous fibres.”
The author has also confirmed the above-mentioned formative process of cutis fibres in the marginal region of the developing operculum (Fig. 6, 8).
The arrangement of cutis fibres in the early stages of development should be regarded as the so-called “growth structure” after KASSOWITZ, which exactly corresponds to the “functional structure” of the cutis in later stages.
The formation of cutis fibres has been thus confirmed to have nothing to do with cellular elements of epidermal tissue, as far as viewed from the morphological standpoint.
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