The physiological functions of the organism and all the phenomena of life are expressed on a cellular basis. Therefore, cytology is essential to the understanding of the genetics, development, and physiological functions of the organism, as well as to pathology and pharmacology, which are the basis of medical treatment, and to breeding in agriculture.
Foundation Stage
The vast majority of cells are very tiny, beyond the limits of human vision. A microscope must be used to observe cells. However, until the objective existence of cells was recognized, it was not possible to know that the objects observed under the microscope were cells. Therefore, in 1677, A. van Levenhuk used a simple microscope of his own making to observe the "spermatozoa" of an animal, but he did not know that it was a cell. Cell (cell, from the Latin cella original meaning space, small chamber) the word is 1667 R. Hooker in the observation of cork slices see cork contains a small chamber and named after it. In fact, these small chambers are not living structures, but the cell walls of the empty space, but the term cell has been used. In the enlightenment period of cytology, although many small objects - such as bacteria, ciliates, etc. - were observed with a simple microscope, the purpose was mainly to observe some developmental phenomena, such as the metamorphosis of butterflies and the structure of sperms and eggs. Due to the limitations of the microscopes of the time, the observations were not precise enough, and the constraints of religious beliefs, the results of these observations instead supported the dogma of preformationism. Some claimed to have seen specific, microscopic "little people" in the sperm, which they believed developed into future individuals - spermatologists; others believed that the "little people" existed in the egg - ovozoologists. The influence of the preformation theory lasted for more than 100 years, preventing people from further understanding of the cell on the basis of R. Hooker, and it was not until K. M. Bell discovered the mammalian egg in 1827 that serious observation of the cell itself began. The development of the achromatic objective lens around this time, the introduction of magenta (carmine) and hematoxylin as dyes for coloring the nucleus of the cell, and the beginnings of the sectioning machine and sectioning techniques all created favorable conditions for finer observation of the cell.
The men who gave a great impetus to the study of cells were M. J. Schleiden and T. A. H. Schwann. The former, in 1838, described the cell as being produced by a crystallization-like process in a mucus-like matrix, and first produced a nucleus (and also found a nucleolus). He also regarded the plant as a ****some of cells, as if it were a community of hydrozoans. Inspired by him, Schwann was convinced that both animals and plants were composed of cells. He accumulated a large number of facts pointing out the consistency of the two in structure and growth, and in 1839 formulated the doctrine of the cell. At the same time, the Czech animal physiologist J.E. Pukeno put forward the concept of protoplasm; the German zoologist C.T.E. von Siebold (1845) concluded that protozoa were all unicellular. German pathologist R.C. Filshaw (1855) in the study of connective tissue on the basis of "all cells from the cell", and the creation of cellular pathology. The German zoologist M. Schulze defined the cell in 1861: "A cell is a mass of protoplasm with all the characteristics of life, in which the nucleus is situated."
This above stage can be considered as the foundation stage of cytology. The further development of cytology began with a deeper understanding of the structure of the cell. For it is necessary to have a proper understanding of the structures in order to proceed to their functions.
Study of Morphological Structures
From the middle of the 19th century to the beginning of the 20th century, the study of cell structure, especially the nucleus, made great progress.
The German botanist E.A. Strasbourg first described the coloring objects in plant cells in 1875 and concluded that plants of the same species each had a certain number of coloring objects; in 1885, the German scholar C. Laboure put forward the law that the number of coloring objects was constant; in 1880, Baranetsky described the helical structure of the coloring objects, and in the following year, Pfetzner discovered chromatophores. It was not until 1888 that W. Wardell formally named the coloring objects in the nucleus as chromosomes. The German scholar H. Henkin observed the X chromosome in the sperm cells of insects in 1891, and the Y chromosome was discovered in 1902 by W.L. Stevens, E.B. Wilson and others.
The phenomenon of cell division, by that time, had been taken seriously and carefully analyzed. The German botanist W. Hoffmeister in plants in 1867, and A. Schneider in animals in 1873, described indirect division in some detail; the German cytologist W. Fleming in 1882, after discovering the longitudinal division of the chromosomes, proposed the name mitosis in place of indirect division, and E. Hoyzel described the distribution of chromosomes during indirect division; after him, E.A. Strasbourg divided mitosis into prophase, metaphase, prophase, and prophase, which have been common until now; he and other scholars also observed meiosis in plants, and after further study finally distinguished between haploid and diploid chromosome numbers.
The study of the cytoplasm is far less thorough than that of the nucleus. Although the German biologist O. Hertwig discovered the centrosome in 1875, it was only through later studies of mitosis that he gained a more detailed understanding of its evolution during mitosis. As for the structure discovered by Golgi (1895), which he called Apparato reticulare interno (later called Golgi apparatus), there was a controversy whether it existed or not before the introduction of the electron microscope. Because this structure can only be seen after the cells have been fixed with a certain fixative and stained with silver or osmium acid, it was thought to be an artificial artifact; however, it can certainly be seen in certain positions of the secretory cells when observing living cells or when staining them with live stains or frozen sections. With regard to the mitochondrion, since its discovery and naming by C. Benda in 1897, there has been more agreement as to its existence. In some cells after fixation with certain fixatives, they can be stained with certain dyes and can also be observed in vivo. But under the light microscope its shape is various, either linear or granular or a string of particles; as to whether it exists in the various cells of animals or all living organisms, at that time there is no conclusive evidence.
The cytoplasm itself was even less well understood. There had been various theories, but none of them reflected the real situation. For example, C. Frohman in 1865 thought that it contained fibrous material interwoven into a framework or mesh, and W. Fleming in 1882 wrongly extrapolated the mitochondria, spindle filaments, and other fibrous formations seen in fixed samples to suggest that the cytoplasm was composed of these filamentous components buried in the matrix. The German histologist R. Altmann in 1886 went so far as to suggest that certain small granules were the simplest, living, "elementary organisms of the cell" and constituted the cell because of their special manner of agglomeration; this may also have been due to the misidentification of the mitochondria as well as the secretory and storage granules. More readily accepted is the honeycomb or foam doctrine of the German zoologist O. Beechley of 1888: the cytoplasm is composed of fine honeycomb-like structures formed by a stickier substance (hyaline hyalopla-sm) filled with another substance called cytosol (enchylema). This doctrine corresponds to a certain extent with reality, for Beechley based his proposal not on the observation of fixed specimens, but on the observation of protozoa in vivo. The cytoplasm of protozoan sunworms is indeed foamy -- the question whether protozoa are unicellular or not was debated for almost half a century, and was not affirmed until 1875 by Beechley's study of the ciliates -- so that the foamy doctrine was maintained for the longest time.
There are two other cases to be traced in regard to the structure of the cytoplasm. In 1899 Garnier, in his study of various types of glandular cells, found that the cytoplasm contained basophilic filamentous or rod-like structures showing dynamic changes, and thought that these were not inclusions in the cytoplasm but were constituent parts of the cytoplasm, thus naming it kinetoplasm, and gave a detailed account of it. This was the cytoplasmic structure, endoplasmic reticulum, which was proved to be true under the electron microscope half a century later, only that it did not receive the attention it deserved at that time.The detailed description of the sarcoplasmic reticulum of the rhabdomyosarcoma of different animals by Verrat in 1902 was also long forgotten, and it was not until after the application of the electron microscope that the accuracy of his observations was fully evaluated in 1960.
Knowledge of cytoplasmic structure lagged behind that of the nucleus or chromosomes, and this situation did not improve for a long time. Especially after the early 1900s, chromosomes were better understood as cytogenetics studied the chromosomal basis of genetic phenomena such as segregation, recombination, interlocking, and exchange.Multilineage chromosomes were discovered in the cells of the Malpighian tubules of mosquitoes by H. Bauer in 1933, and such constructs were found in Drosophila by T. S. Painter in 1934, and in Anopheles shakespeare in 1934 by R. L. King and H. W. Beams. The multilineage chromosome is a huge chromosome found in certain glandular cells of the larvae of Diptera, and in Drosophila it is about 100 times as long as the normal chromosome, each chromosome consisting of many (up to 400) stained fibers, showing y stained banded regions and lightly stained inter-banded regions throughout the length of the chromosome. Its formation is due to intranuclear mitosis (where only the chromosomes divide and the nucleus does not), thus each multilineage chromosome is actually formed from many chromosomes. The large size of this chromosome facilitates analysis of the fine structure of the chromosome. In addition, it is possible to determine the functional activity of a multilineage chromosome based on the distended vesicles on the chromosome. At the same time, however, there was not much progress in the understanding of the structure of the cytoplasm, except for the understanding of some of its physiological functions in conjunction with cell physiology. This situation did not change until the 1940s, when the electron microscope was widely used and a set of techniques for embedding and sectioning specimens was gradually perfected. Through a lot of work, not only figured out the former in the light microscope can be seen but can not see, or still controversial organelles, such as mitochondria, Golgi apparatus, centrosomes, the endoplasmic reticulum, cilia, flagella, and other structures, and also found a lot of previously unseen structures, such as lysosomes, peroxisomes, ribosomes, constituting a variety of cytoskeletal fibers, as well as high-voltage electron microscopy observed by 1 ~ 10? thick and thin fibers consisting of The microbeam system supporting various organelles was observed by high-voltage electron microscopy, especially the various membranes of the cell. In the past, we had never seen cell membranes or nuclear membranes under the light microscope, and had only judged their existence by the interface or physiological conditions, but under the electron microscope, we determined that all membranes were three-layered structures 75 to 100 angstroms thick (called unit membranes). Not only that, but all parts of a cell's membranes are connected, the plasma membrane to the endoplasmic reticulum, and the endoplasmic reticulum to the Golgi apparatus or nuclear membrane. The nuclear membrane is bilayered, consisting of inner and outer membranes, and has a nuclear pore with a certain structure through which the substances of the cytoplasm and those of the nucleus are exchanged. Intercellular connections are also found in the plasma membrane: bridges, tight junctions and gap junctions. These structures are associated with intercellular bonding or the exchange of material between cells; they can be better visualized using freeze-etching techniques.
In the course of 20 to 30 years, considerable insight was gained into the morphology of the cytoplasm as well as the organelles. Of course, the light microscope remained an indispensable and powerful tool in an era when electron microscopes were widely used. The complete cytoskeleton, for example, was observed under the light microscope using fluorescently labeled immunoantibodies.
During this period, less progress was made in the study of the nucleus. Although the structure of the nucleolus was precisely described, with regard to chromatin, the observation of ultrathin sections with an electron microscope revealed only a few dots of coloring -- presumably sections where the chromatin had been cut off -- and no complete chromatin structure could be seen. When the chromatin was spread out, only fibers of different thicknesses could be seen. It was not until the 1970s that nucleosomes were observed under the electron microscope; shortly thereafter, in combination with biochemical extracts, it was observed that chromosomes in mid-division are centered on the so-called scaffolding proteins, from which the DNA fibers extend out in a circular pattern in all directions. But how the chromatin condenses into chromosomes is difficult to determine, although there are different ideas -- some, for example, suggest that it is due to the fact that the chromatin fibers spiral over and over again (the so-called superhelix), but to what extent this corresponds to reality.
Studies of Function
This aspect of research, which has been driven to a considerable extent by other disciplines, can be roughly divided into several phases according to the influence of the various disciplines, which, of course, are not separable from each other.
Embryological Influence In the case of cellular function, it is not possible to find a cell in a mass of tissue as an object of study, as in the case of structure. The egg is a cell, and in an age when individual cells could not be obtained for study, it was extremely convenient material to utilize. Since the egg is used, the study of its various parts must of course be judged by their effect on development. This involves the problem of embryology. But if the function of the heterozygous sperm nucleus is studied by crossbreeding, it is necessary to judge it on the basis of the appearance of heterozygous traits, and this involves the problem of heredity. The early work in this area was essentially carried out by embryologists and was characterized by comprehensive studies, not studying the egg from a purely cellular point of view, but taking the egg as a cell to study problems relating to development, heredity, etc. Some of the major problems have been outlined, and thus have had a profound influence on subsequent academic thought. Brothers O. Hertwig and R. von Hertwig first saw the fertilization of the living egg in 1887, using the sea urchin as a material, and experimentally analyzed fertilization. If the roles of the cytoplasm and the nucleus in development are considered separately, T. H. Boveri's analysis of the phenomenon of chromatin ablation found in the horse roundworm proves that the factors affecting ablation are present in the cytoplasm. In addition, the work on cellular genealogy, in which the oocytes are numbered in order to trace the ins and outs of each oocyte, and the study of the type of cleavage of various types of eggs with varying yolk content, point to the fact that the distribution of the cytoplasm in the oocyte influences the direction of the spindle, determines the formation of cleavage surfaces, and determines the type of oocyte cleavage. Not only this, but in some particularly suitable eggs it can be seen that the material for the formation of various organs is already laid out in the egg, and that after the cleavage of the egg the individual cleavage spheres have a certain correspondence with the organs that are to be formed. All this suggests that the nuclei are equivalent in hereditary potential, and are regulated differently only in later development, through cytoplasmic or intercellular interactions.
The role of the nucleus has also been well evaluated. In 1887 the German experimental embryologist, T. H. Boveri, fertilized sea urchin eggs by two spermatozoa, and, on the basis of the distribution of chromosomes in each ooglobe and the development of the individual ooglobes, concluded that each chromosome was qualitatively different and that the chromosomes were individualized. Using sea urchin eggs, T.H. Morgan accomplished artificial parthenogenesis in 1896 - the development of eggs without fertilization. Fertilization of egg masses without nucleus or with heterozygous spermatozoa, and study of the respective roles of cytoplasm and nucleus in development, it was observed that the resulting larvae all showed the characteristics of the father. All of these demonstrated the importance of the nucleus. Summarizing the achievements of the time, in 1883 the German embryologist W. Roux once expressed the conception that "not only the chromosomes, but also the various parts of each chromosome may be important in determining the development, physiology and morphology of the individual." In 1887 the German zoologist A. Weismann formulated the hypothesis of germplasm. Although this hypothesis was disproved by later experimental studies, there are certain ideological links to be found between the determinants proposed in the hypothesis and later genes.
Removing the influence of academic ideas, an important experimental method was also provided for cytology in order to solve the problems of embryology, which was tissue culture. The American embryologist R.G. Harrison created the method of in vitro culture in 1907 to study the growth of nerve fibers, which was later taken over by the American physiologist A. Carrell and developed into a specialized technique. after the 1930's it showed its importance more and more, and today it is an indispensable technique not only for the study of living cells in all its aspects, but also for many other disciplines.
Influence of genetics After the rediscovery of G.J. Mendel's research achievements in 1900, the study of genetics gave a strong impetus to the progress of cytology. American geneticist and embryologist T.H. Morgan studied the genetics of Drosophila and found that the occasional white-eyed individuals were always males; combined with the existing, knowledge of the sex chromosomes, explaining the emergence of white-eyed males, began to explain the phenomenon of heredity from the cell, the genetic factor may be located on the chromosomes. Cytology and genetics were linked, and quantitative and physiological concepts were obtained from genetics, and qualitative, material and narrative concepts from cytology, gradually giving rise to cytogenetics.
In 1920 the American cytologist W.S. Sutton further pointed out the parallelism between genetic factors and chromosomal behavior, which necessarily implies that genetic factors are located on chromosomes, and mentioned that if two pairs of factors are located on the same chromosome they may or may not be inherited in accordance with Mendelian laws, which foreshadowed the concept of chaining and deepened the concepts of maturation and division and especially of chromosome pairing, and chromosome exchange.
In addition, the discovery of the phenomenon of radiation (X-rays, radium radiation, ultraviolet rays) and the ability of temperature to cause mutations in Drosophila made the experimental study of chromosomes more favorable because of the high frequency of mutations. Various mutations caused by radiation, including gene displacement, inversion and deletion, can be found in the chromosome. Using mutant and wild-type crosses and statistically processing their progeny, the genetic arrangement of the chromosomes can be deduced.
The discovery of multilinear chromosomes opened up new avenues of chromosome research. After concluding that the multilineage chromosome is a bold, paired chromosome, on the one hand, the structure of its detailed study, found that the chromatophores on the chromatophore, many neighboring chromatophores aggregated into a banded area, the chromatophore is not easy to see clearly, but if the staining of the appropriate or in the ultraviolet light can be seen that they are not arranged in a straight and parallel, but a very loose spiral. On the other hand, it is possible to match the gene arrangement on the chromosomes deduced from the linkage groups by the so-called salivary gland method with the morphological chromosome diagram; the hybridization experiments and the morphological observation of the cells can be perfectly corroborated with each other, and it is possible to see the gene arrangement on the multilineage chromosomes in a much more concrete and exact way, and each banded region actually contains more than one gene. Moreover, some mutations are due to positional effects of genes, such as the bar-eye mutation, which was first evidenced on multilinear chromosomes.
Driven by the search for the material basis of heredity, the study of chromosomes has been carried out on the surface, not only for genetic research material, but also many other animal and plant species (some counted about 12,000 species of vascular plants and more than 500 species of mammals), cell division (meiosis), chromosome behavior, chromosome mapping have been studied. Species in the same genus tend to have the same number of chromosomes; however, species in the same family either have unequal numbers or this genus has multiples of another (polyploidy). Individual chromosomes of the same body may not appear to be very different at a cursory glance, but on closer examination they are, so it is possible to state precisely the number and shape of the chromosomes of a species, as well as the size of the individual chromosomes, and to number them in a lineup. It is possible to compare the chromosomes of closely related species, and thus to search for evolutionary relationships; the study of karyotypes points out that the number of chromosomes in similar species may be exactly the same, but there may also be very marked differences, and in the latter case the original form, and the various forms derived from it, can always be found after careful study. In plants three kinds of mutations have been known: polyploidy, the breaking of a chromosome into several small ones or, on the contrary, the assembling of several small chromosomes into a single large one, and the doubling of a pair of chromosomes. These three mutations are sometimes associated with the formation of subspecies and species. In addition, the study of polyploidy in plants has led to the use of various methods, such as chemicals, temperature, and radiation, to induce the production of polyploidy, which has gained value in application in some plants.
Widespread research on the morphology of sex chromosomes has also found a cytological basis for the determination of male and female sex. Some animals are XX and XY, some are ZZ and ZW.
The influence of cellular physiology , the study of the function of other parts of the cell by experimental methods at this stage did not give satisfactory results. With the microscope can not observe the cell membrane, can only be based on the exchange of substances between the cytoplasm and the outside world to determine its existence, as well as the permeability of certain substances, in order to determine some of its functions. Since it is generally said that fat-soluble substances can easily enter the cell, it has been hypothesized that the cell membrane may be composed of lipids or pores of lipids. It has also been hypothesized that the cell membrane is like a filter layer, with small pores that prevent large molecules from entering the cell, because of the difference in the ease of entry of substances of different molecular weights -- the larger the molecular weight, the more difficult it is to enter. In addition, the charge hypothesis has been proposed to explain the extremely complex process of cell permeability based on the fact that electrolytes, such as cationic and anionic ions, permeate cells, and that the acidity of the cellular environment can affect, and thus change, the permeability of cationic and anionic ions. As for the phagocytosis of solid particles, simulation experiments, such as the phagocytosis of chloroform droplets by amoebas, suggest that this is due to the fact that the cell has greater adhesion to the surface of a foreign body than to its surroundings, and that the adhesion causes a local change in the surface tension of the cell membrane, resulting in the swallowing of the foreign body.
These assumptions, even then, seemed to suggest that the cell membrane was passive in terms of permeability; but the cell was also capable of actively ingesting or excreting certain substances against a diffusion gradient or a concentration gradient. Thus it was also envisioned that there might be energy-requiring processes in the cell membrane that were significant for these processes, but the information was not available at that time.
The understanding of cellular respiration at that time was mainly limited to the production of heat from food through the action of various enzymes. Knowledge of the several enzymes involved in this process, such as certain dehydrogenases, oxidases, cytochromes a, c, b, etc., led to the understanding that the combustion of food in the cell does not release all the energy in the form of heat by a single, sudden oxidation, but that it is a gradual process of obtaining and utilizing a small amount of energy step by step through a series of small stages. This process is made possible by the addition of many enzymes to the total respiratory process as transferring oxygen, accepting hydrogen, redox systems, etc., and is finely regulated.
The Influence of Other Disciplines In the early 1940s, technical methods from other disciplines were successively used in the study of cytology, opening up new horizons and forming a number of new fields. The first was the application of the electron microscope which gave rise to ultramicro-morphology.
The Belgian zoologist J. Brashear started from the problems of embryology and used specialized staining methods (Unna, Feulgen) to study the significance of nucleic acids in development. At about the same time, the Swedish biochemist T.O. Caspersson created the ultraviolet cell spectrophotometer to detect the presence of the substances proteins, DNA and RNA in cells, based on the absorption of various substances at certain wavelengths. If the former could be qualitative based on staining, the latter could be quantitative based on absorption. In essence, it was their work that drew attention to the role of nucleic acids in cell growth and differentiation. On the basis of their work, cytochemistry was developed, the study of the chemical composition of the cell, which could be complemented by studies of morphology, adding to the understanding of cell structure.
Analysis of chromosomes with multiple lines, photographed under ultraviolet light, showed that chromatophores as well as nucleoli contained DNA, in contrast to chromatophores, which contained little or no DNA. Digestion with proteases (which may be impure) can dissolve them, and it has been mistakenly assumed that the chromatids are made of protein. In addition, the percentage of certain amino acids in chromosomal segments (euchromatin and heterochromatin) can be accurately determined from ultraviolet absorption spectroscopy. The euchromatin segments appear to contain more high-molecular-weight globulin-type proteins, whereas the heterochromatin segments contain more low-molecular-weight histone-type proteins.
Biochemical cytology arose in the 1940s from the gradual development of biochemical studies of the function of cell parts. Firstly, homogenization - mechanical grinding of cells in a suitable solution - and differential centrifugation were used to obtain mitochondria, microsomes and thylakoids in addition to the nucleus. They were studied separately to understand the presence and distribution of some substances and enzymes, as well as where certain metabolic processes take place. Mitochondria have been more successfully isolated because their size has been measured by electron microscopy and a rough idea of the biochemical processes taking place in this organelle has been obtained, recognizing their importance for energy metabolism. Mitochondria were once mistakenly thought to be an organelle. It was later learned that it was a product of the isolation conditions of the time and was a complex consisting of ribosomes and a small amount of endoplasmic reticulum. Nevertheless, studies such as this one on mitochondria and microsomes have pointed out that many basic biochemical processes take place in the cytoplasm rather than in the nucleus. Such an approach combined with in-depth morphological studies has led to an increasingly profound understanding of processes in the cell.
The use of radioisotopes has opened new avenues for studying metabolic processes in cells. From their participation it is possible to trace precisely the synthesis and transport of substances in the cell, as well as the utilization of stores. For example, it has been shown that phosphorus compounds are incorporated not during mitosis, but during interphase, shortly before the onset of division, and are then distributed to the nuclei of daughter cells. From some of these and other results obtained with other isotopes, it is possible to infer the operation of some important substances in the cell.
While tissue culture was developed to a large extent in the 1930s, only blocks of tissue could be cultured, not yet the individual cells of normal tissues, and it has not yet fully demonstrated its importance. The use of cultured cells allows for the study of many problems that cannot be studied in the whole (in situ), such as cell nutrition, motility, behavior, and intercellular relationships. Almost all kinds of tissues, including some invertebrates (cuttlefish, sea squirts, fruit flies, etc.), have been cultured. Various types of cells that grow from tissue blocks under good culture conditions grow differently. Morphologically they can be divided into essentially three types, epithelial, connective tissue, and wandering cells (e.g., lymphocytes, monocytes, and macrophages). Sometimes cultured cells display characteristics that normal tissues do not exhibit in the organism, e.g., cells from a variety of tissues can acquire phagocytosis if the medium contains substances that enhance surface activity. However, they retain their unique properties and potential because they can grow as they did if the culture environment is changed or if they are moved back to their original site in the animal.
It is worth noting that the growth of fibroblasts in culture is also influenced by the substrate. In general they grow radially and aimlessly from the tissue mass. But if the medium is artificially brought under tension in a certain direction, or if traces are artificially made in the substrate, the cells grow out in the direction of the tension or along the traces. This phenomenon may be used to explain the functional adaptation of connective tissue and tendons in general -- they always grow and differentiate in the direction of tension.
It can be seen that the study of the cell, the use of the electron microscope in the sub-microscopic structure of the depth, and the application of biochemical technology in the function of the depth, has been in the cell biology - in the molecular level of the study of the cell's life phenomena - the formation of the conditions. So later, under the influence of molecular genetics and molecular biology excellent achievements, the cell biology of this new discipline quickly formed.