Acute plaques in multiple sclerosis, their pathogenic significance and the role of spirochaetes as etiological factor.
by Gabriel Steiner, M.D.
Journal of Neuropathology, 11:343-72. 1952.
“In this respect the discovered spirochetes were totally different from the treponema-type and resembled the borrelia-type of spirochetes.”
In the chronic form [of multiple sclerosis] the findings at the time of death represent a terminal cross section of the course of the disease; previous inflammatory reactions may have disappeared partially or entirely. In many other chronic inflammatory disease entities of the central nervous system and other organs of the human body, acute phases and chronic forms of the same disease entity are of common occurrence.
It is the purpose of this paper to contribute some observations to the study of subacute cases of multiple sclerosis, to discuss some of the pathogenetic features of acute plaques in multiple sclerosis and to establish a relationship between spirochetes and tissue reactions in multiple sclerosis.
...Acute plaques in multiple sclerosis occur, significantly, not only in subacute cases of several months' duration but also in older cases in acute clinical relapse. Equally and especially important is the presence of older plaques in subacute cases, indicating morphological manifestations dating back to clinically latent initial stages of the disease at its earliest onset. With these findings in mind, separation of multiple sclerosis into two distinct and different entities seems not justified. [some of Steiner's peers had argued that there were two disease entities -- one an acute disease, the other a purely degenerative chronic disease]
The search for the causative agent in the diseased tissues promises more success when earliest stages of tissue alteration are available for such investigation. Significant pathogenetic information is furnished by the following case of subacute multiple sclerosis.
History: A Negro, aged 30 years, was in good health until May of 1948. At that time he noticed weakness of the left leg. This gradually progressed so that he was forced to drag his left leg, and to remain home from work. The paralysis then extended to the left arm and the left side of the face. His condition grew rapidly worse. Finally he was unable to walk or talk and was admitted to Receiving Hospital on August 7, 1948. ...Course: The patient improved slightly with supportive therapy until August 20, 1948, when his temperature rose to 102 F. Five days later he developed hyperthermia and fell into a deep coma, without recovery. He expired the following day.
...Post Mortem Findings... Central Nervous System: The entire brain and two small pieces of the spinal cord were obtained...There were multiple demyelinated plaques in the right parietal region, mostly in the white matter...In the left hemisphere there were a few scattered plaques also...In sections of the spinal cord no definite plaques were found but some whitish discolorations in wedge shape with the base at the periphery.
Microscopic Observations: In myelin sheath preparations, numerous demyelinated plaques were seen (fig. 1). In these plaques the myelin sheaths were either completely lost or a few islands of preserved myelin sheaths were seen, but the complete loss prevailed. The demyelinated plaques were of varying dimensions. In fat stains these demyelinated areas showed equal distribution of the fatty products; in older plaques, however, these fatty products were lacking or remained in larger quantities only in adventitial spaces of blood vessels. ... The nerve cells in plaques were well preserved (Nissl stain). ...When grey and white matter participated in one single demyelinated plaque the demyelination took place in grey and white matter alike without respect to the border-line between grey and white matter. ...
The Special Silver Salt Reduction Technique to Demonstrate Granular Bodies and Spirochetes:
[a detailed description is provided].
Extracellular Granular Bodies: These granules were of varying sizes and shapes. Round, ovoid, or irregularly contoured shapes were common. ...Two or more granules in close proximity were also seen. The dimensions of these extracellular granules varied from the size of a mast cell granule to one of the size of a red blood corpuscle or even of a small glial nucleus. Often the extracellular granules and astrocytes containing intracellular granules were accumulated around blood vessels in perivascular parenchyma. They were seen abundantly also in the parenchyma without any relationship to blood vessels and less frequently in vessel walls themselves. The photomicrographs (fig. 4) show better than any detailed description the shapes, sizes and locations of these extracellular granular bodies.
[a series of photographs of spirochetes and spirochetal granules found in the autopsied tissues is provided]
Intracellular Granular Bodies: ...The granules differed in shape and size from the extracellular granules. They were more massive, and of a very irregular shape. Nevertheless, they were of the same black, shiny color. ...The intracellular granules were demonstrable by the silver techniques I and II.
Spirochetes: In their fully developed, not yet disintegrating forms, the spirochetes appeared as screw-like organisms. ...Crests and roots were always rounded, never pointed. The minimal thread angle was 60°, its maximum 130°, the average being 97°. In this respect the discovered spirochetes were totally different from the treponema-type and resembled the borrelia-type of spirochetes. ...
Knobs at one end were not unusual (figs. 8d, e, and 9c). There were also loops in the center of the longitudinal axis or more toward the end. The spirochetes were completely detached from any tissue elements. ... The spirochetes were found in marginal areas of acute plaques and in perifocal location in areas close to the periphery of acute plaques, often in histologically seemingly intact tissues. They were always found in locations where abundant extracellular and intracellular granular bodies were present. This close spatial relationship between the spirochetes and the granular bodies is of the greatest practical importance in finding intact and well preserved spirochetes. When masses of extracellular and intracellular granular bodies are found, spirochetes should be looked for at the peripheral areas of an acute plaque containing granular masses close to and in the normal tissues of the central nervous system. One should not expect, however, to find such enormous masses of well preserved spirochetes, as for example, treponemas are seen in the organs of congenital syphilis... The highest number of individual spirochetes in brain and spinal cord of this polysclerotic case were 8 in a high power oil immersion microscopic field. ...
The not disintegrated spirochetes [8 were found] apparently represent stragglers left behind by an enormous army of regularly coiled individual spirochetes. If the granular bodies seen in abundant masses are remnants of disintegrated spirochetes these microorganisms must have been present in enormous numbers and apparently their individual life-span must have been short, in inverse ratio to the speed of reproduction. Excessive reproductive activity of spirochetes, that is speed of multiplication on one hand and very short life-span of the individual spirochetes on the other, are conclusions to be made from the histological appearances.
Significance of Granular Bodies: The very abundant accumulation of granular bodies in close regional relationship to polysclerotic plaques of acute or subacute order can easily be detected. Thus, the granular bodies were seen by many observers (Austregesilo (son) (11), Steinger (12), Guirand (13a, b), Rogers (14), Austregesilo (father) and Fortes (15), Scheinker (16a,b), Marburg (17). These granular bodies were interpreted differently by the various observers, but all agreed that they represented products of disintegration either of tissue elements (Marburg (17)) or of microorganisms. Only Giraud (13a,b) claims that the bodies are the microorganisms themselves. ...
The Relationship of Granular Bodies and Spirochetes: There are all intermediate stages between well preserved regularly coiled spirochetes and granular bodies. There are terminal granules with adherent spirochetal threads (fig. 9c); there are granules already freed from the still persisting spirochetal thread, but at a very short distance from it, so that the breaking off of the granule from the spirochetal thread seems very probable. ...There are spirochetes...still showing the structural continuity between the granule and the spirochete. The knobs and loops represent probably the earliest transitional phases from the spirochetal form to granule formation. There is no doubt that the granular bodies, the haptocytes and the spirochetes are in intimate pathogenetic relationship. ...
The biological significance of these bodies in multiple sclerosis is still obscure. One aspect, however, is certain: These granular bodies are definitely related to the presence of well preserved spirochetes and their disintegrating forms.
Granular bodies in general may represent 1) involutional forms (a) with possibility of redevelopment into typical spirochetal forms, (b) representing beginning disintegration and final death of the spirochetes, (c) possibility of (a) and (b), that is, redevelopment into spirochetal forms as well as irreversible disintegration; 2) specific evolutional forms in the life-cycle of the spirochete. At present no decision between 1) or 2) is possible. ...Experimentally we can produce the granular bodies in cultures of well known spirochetes by exposure to a temperature to 56° centigrade. ...
In the case of multiple sclerosis it is certainly premature to speculate about the significance of the granular bodies in the life cycle of the specific spirochetes. Nevertheless, the value of the granular bodies as indicators of spirochetal presence in the tissues cannot be underestimated.
In Table I a short review of positive findings [of spirochetes or granules in cases of multiple sclerosis] has been compiled. ...Until 1936, among 48 examined cases of multiple sclerosis, 12 were spirochete-positive (25 per cent). Two of these 12 cases (Brack and Kocheise) showed numerous spirochetes. Silver cells were found in over 90 per cent of the examined cases and haptocytes in 25 per cent. ...
In old chronic and treated cases of general paresis spirochetes are never found and the same is true for old stationary cases of tabes dorsalis. It is well known to the pathologist that the microscopic search for the agent in chronic infections, such as tuberculosis and syphilis is often troublesome and does not succeed. Why should it be different in multiple sclerosis?
EVIDENCE OF SPIROCHETAL NATURE
1. The specific morphology of the spirochetes could be described, photographed, and measured. The spirochetes were seen in large numbers in 4 cases of multiple sclerosis in identical morphology. Nothing like it was found in numerous control cases.
2. The order of magnitude is that of spirochetal organisms. The spirochetal structures are of varying length, from 4-18 microns. Their cross section is round like in all spirochetes. Length, thickness and shape of the spirochetal structures vary in the same range as in other well-known spirochetes. There are, however, enough specific morphologic peculiarities of these spirochetes which distinguish these organisms from other known spirochetes.
3. The staining reactions were the same as shown by other well known spirochetes contained in tissues. The typical silver affinity of all well known spirochetes which is essential for the demonstration of every type of spirochetes in tissues is a definite quality of the spirochetal structures in cases of multiple sclerosis. The specific argyrophilia of the spirochetal surface is common to bacterial surfaces but not to protozoan surfaces. All these bacterial and spirochetal organisms are without the colloidal protection against the silver mirror formation or at least without the same protection as most tissue elements and protozoa possess. That is the reason why tissue structures stain in yellow tinges while bacteria and spirochetes take the metallic mirror-like black silver coating. There is also a difference between the surface of bacterial rods and cocci, as well as of fungi on the one hand and that of spirochetes on the other, insofar as most bacteria and fungi need less time of exposure to silver salts (silver nitrate) to be reduced to metallic silver. A difference between the surfaces of bacterial and spirochetal organisms seems also to be indicated by the fact that the color reaction in living bacterial cultures by reduction of colorless tetrazolium salts (see article of Smith (39)) is quite common while living leptospira cultures at our disposal did not change their natural color into blue or red.
4. The spirochetal structures are completely detached from any tissue elements at both ends as well as on all sides. They appear as foreign bodies in the tissues. Very often an empty space or microvacuole is surrounding the black, mirror-like structure of the spirochete.
5. The spirochetes are seen in the parenchyma of brain and spinal cord itself, as well as in the vascular walls. This also excludes the possibility that these spirochetal structures may be elements of the parenchyma proper or reticular fibrils of the vascular walls.
To the eye of the experienced microscopist the reported structures cannot be explained otherwise than as specific spirochetes. They differ from any other known type of spirochetes, especially from the treponemas or leptospiras. They resemble the species of the Borrelia group of spirochetes. I named the organisms spirocheta myelophthora, that is, the myelin sheath destroying spirochete.
HOW TO FIND THE SPIROCHETES
The spatial and temporal conditions for a successful search for spirochetes have to be considered first. Not every polysclerotic plaque contains spirochetes. Old plaques with only little inflammatory reactions are unfit for the search. Plaques containing massive amounts of catabolic neutral fat are already too far advanced in the disease process to show spirochetes. ...the normal tissue between two neighboring acute plaques is a good place to look for spirochetes.
The examination for spirochetes in silver preparations is greatly facilitated by the presence of extracellular and intracellular (in astrocytes and microglia cells) granular bodies in masses. Extracellular granules seem to be the first manifestations of spirochetal disintegration. A later phase in intracellular. The finding of extracellular granular bodies is the first step for the search of spirochetes. ...The best preserved spirochetes were found at the margins of an area containing the granular bodies in masses and always at the margins of such a field toward the normal tissues and not toward the inside of a demyelinated plaque.
...Compared to the immense masses of granular bodies, spirochetes are less frequently found. The well preserved remaining spirochetes are to be considered, as already mentioned, as stragglers...
...The search for spirochetes in advanced, non- or slowly progressing cases is too tiresome and time consuming to promise a good chance for finding spirochetes.
...There remains the problems of who should look for these organisms. Diligence and patience of the examiner and ample time at his disposal are necessary requirements. The investigator should be acquainted with the various silver stains used for demonstration of tissue elements of human body and especially of the nervous system such as neurofibrils, neuroglia cells etc. Some knowledge of the appearance of spirochetes in tissue sections of the central nervous system and other organs of the body prepared with silver techniques will easily be acquired.
POSSIBLE SOURCES OF ERROR
1. Do the spirochete-like structures reported herewith represent pathological or normal tissue elements of the central nervous system and have nothing to do with multiple sclerosis? No structures similar to or identical with the spirochetes have been found in over 250 control cases of diversified diseases other than multiple sclerosis and of normal brains. ...the regularity in form and shape, the limitations at both ends of positive spirochetal findings exclude a mistaken interpretation as argyrophilic senile filaments. ...
2. Artifacts. In one textbook of neurology the spirochetal structures found in multiple sclerosis have been considered as artifacts (Grinker and Bucy (42)).
...Control sections of tissues containing numerous known spirochetes... were used in each batch of staining procedures. These sections serve at the same time as controls for the blackness of the silver-mirror on the spirochetes. Those who regard the spirochetal structures as artifacts are without experience in staining techniques of spirochetes or in the appearance of spirochetes in stained tissue sections. ... To assure a wide range of control of artifacts, numerous pieces of tissue from normal brains or from those affected by diversified diseases were exposed to autolysis, then fixed in formalin, embedded, cut and stained in the same way. No structures like those seen in brains and spinal cords of cases of multiple sclerosis were found. ...
3. The disclosed spirochetes cannot be those of syphilis, post mortem invaders, secondary invaders or contaminants. Spirochetes are found in normal man in the mouth, in the respiratory tract, in human feces and in genital regions. They are, for the most part, harmless non-pathogenic commensals. Spirochetes were never found in the central nervous system of normal man.
...[in] the case presented here and previous cases of multiple sclerosis in which spirochetes were found (Steiner (18, c, d, e), (Rogers (14), Austregesilo and Fortes (15), Scheinker (16)) syphilis could be excluded. Moreover, the disclosed spirochetes do not resemble at all the treponema pallidum; they are specific spirochetes, possibly belonging to the borrelia group
It is known that in the terminal phase of life, the last hours before death and after death central nervous system tissues can be invaded by saprophytic micro-organisms. ...The characteristic and typical finding in such cases is the lack of all inflammatory tissue reactions in spite of the presence of masses of bacteria. On the other hand, in multiple sclerosis there are often massive inflammatory reactions at the borders of recent demyelinated plaques, inside of these and their near extrafocal vicinity. These inflammatory reactions are only indirect evidence of past and present activity of the agent. The immediate proof of its activity is its presence in forms of varying stages of well preserved
to disintegrating forms, from regular spirochetal forms, more or less straight forms to all types of granular spirochetal disintegration, extra- and intracellularly. Particularly, the intracellular astrocytic and microglial ingestion of granular bodies is proof of the intravital activity of the disease process and its agents. Nothing like it is seen in other diseases when secondary post mortem invasion has taken place. ...
4. The possibility that the disclosed spirochetes are harmless intravital saprophytes must be excluded also. The histological reactions are evidence of tissue damage and besides no visible microbial saprophytes have ever been found in brains of normal or diseased human beings. ...
5. Another possibility has to be considered: that is the presence of the spirocheta myelophthora as a microscopically visible and detectable agent together with an ultravisible viral agent. This eventuality cannot be ruled out by our morphological studies with the light microscope. However, such a possibility does not detract anything from the etiological and pathogenetic significance of the findings of spirochetes. In principle, it seems to me very precarious to conclude from our inability to find visible organisms that the agent must be a virus. We know at least one spirochetal infectious disease which was considered due to a virus until Gsell (44) established, beyond doubt, its spirochetal origin (Leptospirosis pomona).
6. A personal error in interpretation of the findings has also to be excluded. ...The slides containing spirochetes were shown to Drs. Brines, Olsen, Weller, Wohlwill and Zimmerman. In critical appraisal they all acknowledged the shown structures as spirochetes of a type different from treponema pallidum.
As an argument against the spirochetal findings and their significance it has been said that reproduction of the same findings by others is lacking. In fact, it is shown in Table I that among a total of 31 cases examined by various investigators 11 showed spirochetes (over 35 per cent). Thus, the demonstration of specific spirochetes has been possible in more cases of multiple sclerosis and by other investigators. I, myself, was able to find the spirochetes in considerable numbers in 4 cases (once in brain and particularly in spinal cord, once in a case with a second acute relapse after 12 1/2 years remission, once in a case with miliary granulomas, the fourth case is being reported herein). In cases of chronic type the finding of spirochetes is too sporadic to spend much time and effort. Rogers (14), using Marburg's material in Vienna, found the spirochetes in 1 among 11 cases. Blackman (20) found structures resembling spirochetes in 5 of 11 cases of multiple sclerosis. Scheinker (16b) saw spirochetes in 4 out of 8 examined cases. Austregesilo and Fortes reported 1 positive case of a 38 year old woman with a duration of 8 months of disease. ... Spirochetes cannot be expected to be seen in every case of multiple sclerosis. The best chance for finding the spirochetes is in polysymptomatic cases of short duration (3 to 8 months) or older cases in recent relapses. Old burned-out cases do not offer much chance. ...Without the use of adequate silver methods and without a diligent search a reproduction of my findings is impossible. The time applied for the search is proportional to the success of finding spirochetes. ...
A great number of problems are still unsolved:
How do the spirochetes enter the human body? After the entry of the organisms into the body are there systemic reactions or not? How much time elapses from entry into the body until the organisms reach the central nervous system? Where do the spirochetes harbor in the clinically latent initial phase of the disease? And where do they reproduce? What is the stimulus for a new reproductive phase of spirochetes responsible for relapses and acute exacerbations? ... Are there immunological reactions to the presence of spirochetes in the body?
... A special problem arises when we consider the pathogenic role of the spirochetes and the granular bodies, derived from the spirochetes. Is the activity of motion of these corkscrew-like organisms the essential tissue damaging and myelin-sheath destroying factor? In other words, is a micromechanical injury the real tissue damaging effect? Or are substances of the spirochetes or derivatives of these, for example the granular bodies and their substances biochemically noxious myelolytic agents? Or is the defense reaction of the central nervous system itself, especially astrocytic proliferation and haptocyte formation more harmful and injurious to the tissues, particularly to the very sensitive myelin sheaths than the spirochetes themselves and their granular bodies? In multiple sclerosis the detectable tissue damage may be tardy in its development and quite some time behind the appearance of actively motile spirochetes and their granular bodies. These visible tissue reactions may appear some time after the spirochetes and their granular bodies for the most part have already disappeared. This would not be unusual when compared with other chronic infectious diseases. It could explain also the difficulties in detecting the causative agents in the majority of chronic cases and the good chance of finding the spirochetes in subacute cases or in acute relapses of older cases.
With a new discovery a great number of new problems arise and new ways of investigation will lead to new endeavors which may successfully solve the overall important therapeutic problem of multiple sclerosis.