Art. XII.—On the Production of Inflammatory Action in detached Portions of dead Animal Bodies.
[Read before the Westland Institute, 1st November, 1881.]
The question, “What is death?” is one not so easily answered as might be supposed. The popular idea that death in animals is a sudden and instantaneous change, is of couurse not held by physiologists, who have long recognized the distinction between somatic death, or that of the animal body as a whole, and molecular death, or that of the elementary structures of which it is built up. It is difficult to find a good and terse definition of death. In the following article, somatic death may be defined as the permanent arrest of all the functions and powers of the body. The only certain proof of death, as thus defined, is the commencement of chemical decomposition in the whole of the body
It will be at once apparent that this definition of death leaves a considerable interval between that cessation of respiration and circulation, accompanied by entire unconsciousness, which is the popular idea of death, and the commencement of chemical decomposition. During this interval the only vital actions* generally supposed to continue, are that peculiar state of muscular action called the rigor mortis, and the growth of the hair. Some writers consider even the latter as not a true growth, but only an appearance produced by the shrinking of the skin.
It is the purpose of this paper to give a very brief epitome of a series of experiments which have been performed during the last ten years, showing that, in so far as inflammation may be considered as an evidence of life, molecular life exists with a vigour and for a length of time hitherto unsuspected, after somatic death has taken place.
It may just be mentioned that, when in medical charge of a smallpox hospital some ten years ago, in the West Indies, I was engaged in microscopical investigations into the growth and development of the variolous vesicle, which were published at the time in the “Medical Times and Gazette,” 1871–2, in a series of papers on the Pathology and Treatment of Smallpox. I then observed that in my quarters, a small wooden building of inch planks, where the temperature in the middle of the day was often 100° F., changes took place in the variolous matter, kept in the ordinary capillary glass tubes. I was thus induced to try the cultivation of variolous
[Footnote] * I use this term for convenience' sake.
matter, and subsequently of vaccine lymph. These experiments were only partially successful for reasons which are now obvious; but they led on to other experiments.
Mainly these were on the growth and development of the red corpuscles in birds. The results of these experiments were published in the Transactions of the New Zealand Institute for 1874. It was shown that by the use of a suitable nutrient fluid—egg-albumen mixed with water—the nucleated red corpuscles could be made to grow and throw off their nuclei, which became developed into round coloured non-nucleated corpuscles, exactly resembling those of mammalia. The same experiments, with a like result, were made with the blood of fishes, particularly with sharks and rays, and subsequently with the blood of reptilia.
During the investigations upon the blood of reptilia, (frogs being selected for convenience), I naturally took the opportunity of observing the phenomena of inflammation, as seen in the web of the frog's foot and the tail of tadpoles. Having for some time entertained grave doubts, on à spriori grounds, as to the possibility of Cohnheim's so-called wandering corpuscles really passing through the walls of the capillaries, I was induced to make some experiments with a view of testing the matter.
For this purpose I took advantage of a few months' residence in New South Wales, not only carefully and repeatedly to examine with the microscope, the phenomena of inflammation in the tails of tadpoles, but also to show, beyond the possibility of doubt, that leucocytes do not, in these animals, wander from the blood vessels, but are formed from pre-existing germs in the solid tissues, either from connective tissue corpuscles or from some other source. A very simple experiment showed this. A small portion of the transparent extremity of a tadpole's tail was cut off, and immersed in a nutrient fluid, (half egg-albumen and half water was found the most useful), and kept at the ordinary temperature of the air, which at that place and season varied from 70° to 90° F., in the house. Control experiments were made by immersing other portions of tails in water, in water impregnated with carbolic acid, and in various other media. The tails immersed in pure water were in a few hours in a state of decomposition, sulphuretted hydrogen was evolved, bacteria and multitudes of animalcules (monads) were formed in the water, the portion of tail was shrivelled, and dead to all intents and purposes. With carbolic acid the tails underwent no change.
The following description, taken from the notes of one experiment, will show the changes that ensue in a few hours:
“March 13th, 1878, 2.30 p.m—Snipped off four tadpoles” tails, placed them in a mixture of egg-albumen and water. Tails sank in mixture.
5 p.m. temp. 90°F. The cut extremity of each piece was cloudy and whitish, as viewed through a lens of 1½ inch focus; vessels looked more marked and prominent than when put in; outline of pieces not so sharp and well marked as those in the water.
“Under the microscope, round nucleated cells were seen projecting from the cut surface; a few of such cells were floating in the nutrient fluid; several dark-coloured cells round, containing numerous nuclei, were seen.
“At 5.30, these were obscured by a cloud of white cells with granular contents.
“14th, 9 a.m.: The little phials in which the tails had been placed showed a white cloudy precipitate, about ¼ inch deep. On microscopical examination, this proved to be granular protoplasm in amorphous masses, but showing faintly a commencement of segregation into cells. In the fluid were floating about (A) innumerable altered red corpuscles; (B) many round cells of a yellowish or fawn colour, containing two or three nuclei; (G) innumerable leucocytes; (D) columnar and other epithetial cells.
“The tails themselves showed the following changes:—1st, all the red corpuscles were gone; none were to be seen in the most transparent parts; 2nd, all the pigment cells were broken up into small portions, irregular in outline, but much rounder and less angular than the normal pigment cells of the tadpole; 3rd, the striped muscular tissue was in a state of incipient fatty degeneration; 4th, over the whole of the skin were crowds of leucocytes covering it, and easily detached.”
This is a fair sample of scores of similar experiments. They proved that leucocytes originated in an inflamed part, and were not brought to it from the blood.
I then tried what could be done with warm-blooded animals, but was for a long time only partially successful, owing to not using a proper nutrient fluid at a sufficiently high temperature. The following experiment with egg-albumen and water is interesting, as showing that even with this the commencement of inflammation could be produced in a warm-blooded animal.
December 23rd, 1878.—Placed portions of the liver, the Peritonœum from the mesentery, and voluntary muscle of a duck, in a mixture of egg-albumen and water, kept in contact with my own body. The Peritonœum was in a separate bottle. Between two and three hours afterwards, there was evident (naked eye) turbidity of the albumen in which the Peritonœum was placed; the membrane appeared swollen and milky. The turbidity of the bottle in which the muscle and liver were placed was not so marked. Both appeared paler.
December 24th, 8 a.m.—Increased turbidity. Peritonœum much swollen and milk-white; the muscle nearly milk-white; the liver a pale yellowish grey.
Peritonœum bottle; under microscope, multitudes of young cells and granules were floating about in the fluid;* Peritonœum granular, with many young, round, nucleated cells sticking all over it. (Bad light; very wet day.)
Liver very friable; quantities of young cells with well marked edges and nuclei, mostly one, some two, a few three. The usual liver cells were dark, with many dark spots, not removable by acetic acid.
Muscle: no cells except a few adherent to it. No trace of striation, but very opaque fine granules (commencing fatty degeneration).
At length it occurred to me that defibrinated blood would be the best nutrient fluid for mammalian tissues. After numerous experiments, many of which were failures owing to the want of suitable apparatus, and the extreme difficulty of keeping the blood at the temperature desired, ranging from 100° to 105° F., the following were found to be the conditions under which inflammation in detached portions of the body could be carried as far as the production of pus.
The blood should be obtained from an animal rapidly killed by violence, not poison. It should be defibrinated by whipping, and the agitation of the blood should be continued until the whole has become bright scarlet, and thoroughly oxyginated.† The parts to be operated on, which should if possible be from an animal of the same species as that from which the blood is taken, are then to be placed in a glass vessel containing a quantity of the blood—the more blood the better; for instance, 4 fluid oz. of blood would be a fair allowance for a sheep's eye. The vessel must be closed to prevent evaporation of the watery parts of the blood. Open vessels are found not to answer, the blood gets thick very soon. This glass vessel must be placed in a water bath, which must be kept at a temperature of 100° to 105° F. This is the most difficult part of the process. Until one has watched a thermometer for some hours, it will hardly be believed, how, with the same degree of applied heat, the temperature of the water bath will vary. As the temperature of the room falls or rises, so the applied heat has to be adjusted, and a few minutes neglect will suffice for the whole
[Footnote] * Subsequent researches made it probable that these were not derived from the Peritonœum itself but from the lymphatic vessels and their contents, contained within the folds of the mesentery.
[Footnote] † The blood always becomes of a dark venous hue after being exposed to the temperature indicated for a few hours.
experiment to be spoiled. E.g.—One night after watching from 8 p.m. until 1 a.m. I turned away for a few minutes to get a cup of coffee. When I had finished I found the thermometer marked 113° F.; half an hour afterwards the blood was black and smelling most offensively. It may be observed that when once molecular death has occurred in the blood and tissues, chemical decomposition proceeds with very great rapidity.
The only absolutely safe plan to prevent the temperature rising too high, is to place a very small tube or bottle in contact with the body of a warm-blooded animal. The warm-blooded animal I found most convenient was myself, but on one or two occasions I used a fowl, tying the tube or bottle under its wing when at roost. I also tried a cat, but cats and fowls are both objectionable—the former have claws, and the latter claws and beaks. The objection to the healthy human body is the low temperature; you can get cell-growth abundantly; you can get fatty degeneration of muscle, but I have not yet succeeded in getting pus, except at a temperature over 100° F. (38 Cent.) Having only produced pus in the chambers of the eye, I have not yet been able to do so except at a higher temperature than the healthy human body affords. I have tried every tissue of the mammalian body, except bone, repeatedly, but the most striking results were with eyes, and as the globe of the eye roughly removed with muscles attached, contains nearly all kinds of tissue except bone and cartilage, it is very convenient. I will briefly describe the changes that follow immersion for from 4 to 12 hours in defibrinated blood of the temperature 100° to 105° F.
I sent an account of these experiments more than a year ago to Professor Flower, F.R.S., and to Mr. R. Brudenell Carter, with specimens put up in carbolized glycerine, of portions of the retinæ and conjunctivæ, and I think other structures of the eye. Since that time I have several times repeated the experiments with the same results. I am now about to try what changing the blood frequently, so as to give fresh supplies of oxygen, will do.
It may be well to mention that the immersion of an eye in defibrinated blood at a temperature of 50° to 60° F., will in about half a hour restore the transparency to the cornea; making it quite bright like the living eye, so as to make out all the structures with an ophthalmoscope.*
Within a period varying from one to two hours, according to the temperature, a fresh mammalian eye will undergo the following changes when immersed in defibrinated blood of a temperature of 100° to 105°. First, the dull opalescent tint of the cornea will disappear and it will become bright and transparent. Then it loses this brightness again, and becomes of a
[Footnote] * This might be utilized for the purpose of taking photographs after death.
dull milky tinge; not the appearance presented after death, but more opaque. It is difficult to describe the difference, but it will be immediately appreciated when seen. Second, after two hours, or thereabouts, some one point near the centre of the cornea becomes obviously whiter and more opaque than the remainder, and looks rougher and more granular; in fact, looks like a recent ulcer in the first stage. Third, during the course of six or eight hours the cornea becomes quite opaque, so that no portion of the pupil or iris can be seen; the epithetial layer peels off in large pieces with great ease; the whole globe becomes flaccid; the sclerotic much softened; the crystalline is slightly less transparent in sheeps' and pigs' eyes, but small eyes may be semi-opaque; the aqueous humour is turbid; the vitreous reddened but transparent; the retina, except round the optic nerve, converted into dirty-looking pus, mixed with the débris of the pigment layer of the choroid. The recti and other muscles are pale; the cut extremity of the optic is reddened. When portions of the epithetial layer of the cornea are detached, they appear the colour of whitey-brown paper when wetted, but are more opaque. Under the microscope, to quote from the notes, “The shreds of fibrinous exudation in the anterior chamber are composed of granular masses faintly marked out into cell-like portions. The epithetial layer of the cornea is a mass of proliferating cells, the nuclei of the flat epithetial being much enlarged and quite round; immense numbers of cells, four or five times the size of a human red corpuscle, and exactly resembling these enlarged nuclei, were floating about in the immediate neighbourhood of these shreds of epithetium. The columnar and spherical cells were all enlarged, and in many the nucleus was showing a tendency to divide.” The deeper layers of the cornea are full of young cells, and the normal structure is often quite obstructed by their number. In addition to the shreds of fibrinous exudation in the aqueous, “it contains numbers of detached leucocytes.”
The crystalline shows, without any staining or preparation, its fibrous structure; the nuclei of the fibres are much enlarged and very distinct. When the crystalline is detached from the eye and immersed directly in blood it assumes a sort of opalescent tint.
The smaller vessels of the choroid have disappeared, the pigment cells are almost all broken up, and in the most advanced stages nothing but pigment granules can be seen.
The retina, except just round the optic, is, as has been said, dissolved into pus and débris; all traces of rods, cones, and nerve cells, have disappeared. The portion adherent to the optic is of a dirty drab colour, and loaded with leucocytes.
In the vitreous itself, numerous leucocytes, appear to exist, but I am not sure that they are formed in the substance of the vitreous, as they may be merely adherent to the portions examined. In the centre of the vitreous a vein may be observed by the naked eye, “it is gorged with red corpuscles, but there were no leucocytes within it, nor surrounding it in greater numbers than were to be found elsewhere.”
The muscles show commencing fatty degeneration of the fibres.
This is a very brief account of some of the changes that occur. Briefly the whole globe may be said to be in the first stage of acute inflammation.
Portions of lung, treated in the same way, show swelling of the pleura; the air cells gorged with leucocytes, and the lung no longer crepitant.
If in any parts of the body subject to this process there are veins filled with blood, and visible to the naked eye, the corpuscles within them become shrunken, and much smaller. All the red corpuscles in the capillaries and smaller vessels disappear. My opinion, derived from hundreds of observations, is that they dissolve when in vessels which contain only one or two rows, furnishing the material for the formation and growth of leucocytes.*
The following experiment was a little variation from the plan usually adopted:
November 5th, 1880.—A kitten was killed by wringing its neck; its abdomen was opened, and about half an ounce of blood from another kitten, not defibrinated, placed in the cavity of the Peritonœum, together with two eyes, a portion of liver, and a portion of lung from the other kitten. The animal was then wrapped in a warm cloth, placed in a Norwegian refrigerator previously heated, together with a bottle of hot water. It was then kept in a warm room for twenty-eight hours, the bottle being refilled with hot water three times.
November 6th, evening.—It was evident from the smell that decomposition had commenced. The kitten was therefore taken out and exposed during a cold night.
November 7th, morning.—The abdomen was opened. It was first noticed that every particle of the blood put in had disappeared from the Peritonœum. The bowels near that part which had been opened were rough and of a bright red colour; particles of what appeared like lymph were attached to the Peritonœum in flakes; they were of a drab colour, and under the microscope were seen to consist of minute granular particles, bacteria and fatty matter; not a trace of red or white corpuscles.
[Footnote] * This is not the place for the indication of this opinion, but I have numerous notes of microscopical examinations, which may at some future time be published in support of the view here maintained.
The piece of liver was soft, of a pale fawn colour. It was not examined microscopically.
The eyes were in the condition described above.
The pericardium was distended with serum, tinged bright red with blood; the wall of the left ventricle contained a black clot.
In a similar experiment performed with puppies, I find it noted that “striped muscular tissue shows nuclei very distinctly; no blood corpuscles amongst the muscular fibres; blood in capillaries has all disappeared.” This is invariably the case. [See note ante.]
In one puppy, when a portion of pericardium had been removed immediately after cessation of respiration, during a condition of entire unconseiousness, but the heart having beat a few times, I find it noted “the chest and membranes wounded appear inflamed; pericardium found clouded with leucocytes.” In this case the puppy was poisoned with hydrocyanic acid.
To conclude. The experiments, of which I have given a very brief account, and which have been carried on with intervals for several years, extend to hundreds of observations. They show that in parts of the body separated from the trunk after somatic death, the phenomena of inflammation as far as the production of pus may be produced by immersion for some hours in defibrinated blood, at a temperature of 100° to 105° F., or in albuminous fluids of the same temperature capable of supplying them for a short time with nourishment. It will be observed that true nutrition does not occur; there is no evidence that the structures grow; they do not assimilate to themselves the elements necessary for their growth or reproduction; they degenerate; they take on a lower form of life, but still it is life after a kind. The highest forms of tissue, like muscular fibre, simply undergo fatty degeneration; cell structures take on a lower form of cell structure without differentiation.
I have given an epitome of some of the observations made without attempting to theorize on them. Should this paper receive any attention, I may pursue the subject, but in this remote corner of the world, without access to libraries, without personal communication with men engaged in similar pursuits, and with most imperfect apparatus, there is not much encouragement to persevere.