Laurens N. Ruben
Wm. R. Kenan Jr. Professor, Emeritus
|
|
Laurens N. RubenWm. R. Kenan Jr. Professor, Emeritus
|
MACROPHAGE FUNCTION |
T CELL FUNCTIONS |
B CELL FUNCTIONS |
ANAMNESIS |
HAPTEIN-SPECIFIC TOLERANCE |
APOPTOSIS |
METAMORPHOSIS |
MACROPHAGE FUNCTION:
1. Phagocyte blockade will inhibit responses to heterologous RBC’s in
the newt. Ruben, L.N., Jonnes, H. and Stack, J. In " Aspects of Developmental
and Comparative Immunology I. ", Ed. Solomon, B.J., pp.171-178. Pergamon
Press, Oxford, U.K. (1981).
2. The newt can be made to respond to soluble antigens, e.g., keyhole limpet
hemocyanin, and helper function can be demonstrated with them, when the antigens
are bound to particles. Ruben, L.N. and Stack, J. Dev. Comp. Immunol.6:491-498
(1982).
3. The peritoneal macrophages of Xenopus will take up less radio-labelled
ovalbulmin and degrade a smaller percent of what they have taken in, than comparable
cells of the mouse. Moreover, newt Mø are unable to take any in, unless
it is presented adsorbed to particles. Thus, the failure of primitive vertebrates
to respond to soluble immunogens is due to a failure of their Mø's to
take up the potential immunogen. Gammie, A. and Ruben, L.N. Cell Immunol.100:577-231
(1986).
4. Antigen-competition can be demonstrated in Xenopus, but not the newt,
when two species of RBC’s are injected sequentially. The cell type responsible
appears to be the macrophage. Ruben, L.N. and Mette, S.A., Cell Immunol. 51:379-389
(1980).
T CELL FUNCTIONS:
CELL-MEDIATED IMMUNITY:
1. Immunologic factors determine the ability of different frog and salamander
organ implants to stimulate the growth capacity of regeneration potent salamander
limb tissues. Ruben, L.N. Amer. Nat. 94:427-434 (1960).
2. Lymphoreticular cancer development in Xenopus laevis , the
South African clawed toad, suppresses normal allograft rejection capacities.
Ruben, L.N. Amer. Zool. 11:229-237 (1971).
3. The cyclical destruction and re-growth in vivo of foci of lymphoreticular
cancer in Xenopus depends on immunologic maturation for its existence
e.g., Ruben, L.N. Dev. Biol. 22: 43-58 (1970).
4. Mature lymphoid tissue implants suppress the development of the allograft
response in larvae of Xenopus differentially. Ruben, L.N., Stevens, J.M
and Kidder, G.M. J. Morph. 138:457-466 (1972).
HELPER FUNCTION:
1. The capacity of responsivity to heterologous SRBC relates to spleen lymphocytic
development. Its cytodynamics in Xenopus laevis are similar to
those in mammals . Kidder, G.M., Ruben, L.N. and Stevens, J.M. J. Embry. Exp.
Morph. 29:78-85 (1973).
2. Carrier dependent and specific helper function can be demonstrated in a "lower"
vertebrate e.g. the newt, Notophthalmus viridescens. Ruben, L.N., Van der Hoven,
A. and Dutton, R.W. Cell Immunol. 6:300-314 (1973).
3. Carrier dependent and specific helper function can also be demonstrated in
the goldfish, Carassius auratus.. Ruben, L.N., Warr, G.W., Decker, J.M. and
Marchalonis, J.J. Cell Immunol. 31:266-283 (1977).
4. Helper function in Xenopus is thymus-related. Gruenewald, D. and Ruben,
L.N., Immunol. 38: 191-194 (1979).
5. T cell stimulating lectins e.g. Con A , but not wheat germ agglutinin, can
substitute for helper function in Xenopus. Clothier, R.H., James, H.S.,
Ruben, L.N. and Balls, M. Immunol. 52:703-709 (1984).
6. Human IL-2 will substitute for helper function in Xenopus. Its capacity
to do this will last for three hours prior to antigen challenge. Since rIL-2
plus carrier priming fail to further enhance an anti-hapten response, they are
likely to be acting on the same cellular population which is maximumized by
one or the other protocol. Ruben, L.N. Immunol. Letters. 13:227-230 (1986).
7. Human IL-1, particulate Con A , but not soluble Con A or soluble or particulate
WGA or human IL-2, will substitute for carrier primed helper function in the
newt. While anti-human IL-2 receptor antibody will bind specifically to freshly
biopsied spleen cells, Il-2 will not compete with this binding. The shared lectin
specificity suggests that T-like cells are responsible for helper function in
this animal which has an "immature" thymus. The lymphokine and Con
A data suggest that while human IL-2 is unable to affect the cells of this species,
Con A and human IL-1 can, perhaps through the stimulation of autologous IL-2
production. Ruben, L.N., Beadling, C., Langeberg, L, Shiigi, S. and Selden,
N. Thymus 11: 213-220 (1988).
8. The Xenopus spleen cells which are PHA activatable with regard to
IL-2 receptor density and which bind rIL-2, are related to helper T cell function
in cytotoxic and humoral immune responses, since it is removed by N-CH3-N-Nitrosourea
(NMU) injection. NMU is selectively lymphotoxic in the toad and specifically
removes these two functions. Panning spleen cells with a mAb to Xenopus
IgM showed that those cells which bear constitutive IL-2 receptors and bind
rIL-2 are equally abundant in the T and B cells populations of freshly biopsied
Xenopus spleens. Langeberg, L., Ruben, L.N., Clothier, R.H., and Shiigi,
S. Immunol. Letters 16: 43-48 (1987).
9. A monoclonal mouse anti-human IL-2 receptor antibody (anti-p55) will bind
receptors on the surface of immunocytes of Xenopus that are only slightly
larger than 55kDa. Ruben, L.N., Langeberg, L., Malley, A., Clothier, R.H., Lee,
R.O. and Shiigi, S. Immunol. Letters 24:117-126 (1990).
10. Norepinephrine (NE) will stimulate helper function and suppressor function
in adult Xenopus. Low dosage immunization will cause the release of NE
in the spleen that will affect helper function positively and transiently, while
high dosage immunization will affect NE release that is sustained and will stimulate
suppressor function. The effect will be prolonged as long as antigen is around.
Clothier, R.H., Ruben, L.N., Johnson, R.O., Parker, K., Greenhalgh, L., Ooi,
E.E., Sovak, M. and Balls, M. Internat. J. Neurosci. 62: 123-140 (1992).
11. Mortality in developing larvae, particularly during the metamorphic period,
can be manipulated by using IL-2 along with an antigen. Thus, it may be possible
to lower the amount of IL-2 used in experimental cancer therapy by adding an
antigen in conjunction with it (Ruben, L.N., De Leon R.T., Johnson, R.O. and
Clothier, R.H. Interleukin-2-induced mortality during metamorphosis of Xenopus
laevis. Immunol. Letts 51:157-161 (1996).
SUPPRESSOR FUNCTION:
1. Thymus-dependent suppressor function can be demonstrated in vitro
with Xenopus laevis. Ruben, L.N., Mette, S.A, Edwards, B.F. and
Cochran, S. Thymus 2:19-25 (1980).
2. Thymic suppressor function is antigen dependent, is only partially antigen-specific
and is non-MHC restricted in Xenopus. Ruben, L.N., Beunafe, A. and Seivert,
D. Thymus 5:13-18 (1982).
3. While thymus suppressor function is not MHC restricted, there are genetic
limits to its capacity. R, L.N., James, H.S., Clothier, R.H. and Balls, M. J.Immunogenetics
11:97-102 (1983).
4. The differential temperature sensitivity of suppressor function to long exposure
times of cold may account for immune response capacities of ectotherms during
the long cold winters in temperate climes. Ruben, L.N., Clothier, R.H., Buenafe,
A., Needham, P., James, H.S. and Balls, M. Thymus 6:143-152 (1984).
5. Suppressor function is compartmentalized in adult Xenopus:inducer
suppressor cells are in the thymus, while inducer and effector suppressors are
in the spleen. Moreover, both functions are sensitive to cyclophosphamide and
are macrophage dependent. Ruben, L.N., Buenafe, A. Oliver, S., Malley, A., Barr,
K and Lukas, D. Immunol. 54:65-70 (1985).
6. Lectins, e.g. wheat germ agglutinin, peanut agglutinin and Con A can initiate
inducer suppressor function in the Xenopus thymus, but only WGA and PNA
can stimulate splenic suppressor inducer function. Ruben, L.N., James, H.S.,
Clothier, R.H., Barr, K. and Balls, M. Thymus 7:161-167 (1985).
7. Suppressor inducer function of the Xenopus thymus, tested in vitro,
is mediated by a soluble factor. Ruben, L.N., Barr, K, Clothier, R.H., Nobis,
C. and Balls, M. Dev. Comp. Immunol. 9:811-818 (1985).
8. Larval thymocytes have suppressor function which will affect adult immunized
splenocytes. However, during metamorphosis this function is impaired. This impairment
is not due to suppressor cells migrating to the spleen, since the spleen is
also lacking in effective suppressor function during metamorphosis. While wheat
germ agglutinin is an effective stimulator of splenic suppressor function in
adults, it fails to stimulate it during the metamorphic period. By using fluorescein-labelled
WGA, we were able to show that this lack of capacity of wheat germ agglutinin
to stimulate splenic suppressor function during metamorphosis was not due to
the absence of binding by the Fl-WGA. Thus, the loss of function by the thymic
suppressor cells did appear to occur as a consequence of the cells leaving for
the spleen, the adult site of suppressor function. Kamali,D., Ruben, L.N. and
Gregg, M., Cell. Diff. 18:225-231 (1986).
9. Corticosteroid regulation of IL-1 and IL-2 appears to be responsible for
deficient immune suppressor function in the thymus during metamorphosis. Highet,
A. and Ruben, L.N., Immunopharm. 13:149-155 (1987).
10. CyP, but not IL-2 will break the tolerance initiated to single haplotype
disparate skin grafts made during metamorphosis that are sustained for at least
100 days. Thus, suppressor function appears to be involved in the maintenance
of a tolerance which is susceptible to rIL-2 injection, while it is being established.
Horton, J.D., Horton, T.L., Varley, C.A. and Ruben, L.N. Transplant. 47:883-887
(1989).
11. There are two types of suppressor cells with respect their differential
sensitivity to CyP. Clothier, R.H., Last, Z., Samauroo, J., Ruben, L.N. and
Balls, M. Devel. Comp. Immunol. 13: 159-166 (1989).
12. Xenopus suppressor factors will bind antibodies raised against inducer
and effector suppressor factors of mouse in Western blots and in ELISA. Moreover,
the inducer factor also binds an anti-IL-10 Ab raised against mouse IL-10. Recently,
we have found that a protocol that generates suppressor inducer factors in Xenopus
will apparently also stimulate mRNA expression particularly of IL-10, IL-5 and
IL-4 genes in Xenopus, using RT-PCR with rat oligoprimers of the cytokines.
RECOGNITION AND BINDING:
1. Antigen-binding receptors of primitive lower vertebrates vary in their capacity
to discriminate among different haptens after hapten-specific challenge., Ruben,
L.N. and Edwards, B.F. Cell Immunol. 31:437-442 (1977).
2. Alpha and ß adrenergics differentially effect the capacity of immunized
B cells to bind antigen. Hodgson, R.M., Clothier, R.H., Ruben, L.N. and Balls,
M. Eur. J. Immunol. 8:348-351 (1978).
3. Amphibian antibody and complement-dependent lysis is temperature dependent.
Ruben, L.N., Edwards, B.F. and Rising, J. Experientia 33:1522-1523 (1977).
4. Immunologic stress may cause the newt to release a low MW antibody which
is normally not produced. Warr, G.W., Ruben, L.N. and Edwards, B.F. Immunol.
Letts. 4:99-102 (1982).
B CELL SUBPOPULATIONS-TI IMMUNOGENIC CARRIERS:
1. Two subpopulations of hapten-specific B cells, which are activated by different
carriers, can be demonstrated in Xenopus. Horton, J.D., Edwards, B.F.,
Ruben, L.N. and Mette, S.A. Dev. Comp. Immunol. 3:621-633 (1979).
2. Two sub-populations of hapten-specific B cells which respond to different
carriers can be demonstrated in the newt. Ruben, L.N.and Reischel,G. Dev. Comp.
Immunol. 5:513-518 (1981).
3. Responsivity to TNP-Ficoll varies with developmental stage and internal endocrine
environmental changes with age. Ruben, L.N., Clothier, R.H., James, H.S. and
Balls, M. Cell. Diff. 14:1-5 (1984).
4. T cell-stimulating lectins, e.g. Con A can substitute for the thymus requirement
for responses to TNP-Ficoll in Xenopus. Clothier, R.H., Ruben, L.N.,
James, H.S. and Balls, M. Immunol.52: 483-489 (1984).
5. Murine and human IL-2 can substitute for the thymus requirement in TNP-Ficoll
responses in adult Xenopus. Ruben, L.N. Clothier, R.H. Balls, M., Cell.
Immunol. 93:229-233 (1985).
6. The TNP-Ficoll response is regulated through two different pathways in Xenopus.
The TNP-Ficoll response is T cell-requiring, but this requirement can be substituted
for by any protocol which stimulates peripheral T cell activity, including allograft
rejection or by optimizing the function of peripheral, but not thymic Mø's.
Clothier, R.H., Ruben, L.N., Smart, C. and Balls, M. Dev. Comp. Immunol. 10:577-583
(1986).
7. Responses to TNP-PVP are very similar to those to TNP-Ficoll, i.e. the two
will compete for B cells, except that the TNP-PVP is less thymus-dependent.Clothier,
R.H., Kandola, L., Mirchandani, M., Ellis, A., Wood, P., Last, Z., Balls, M.
and Ruben, L.N. Cell. Diff. 23:213-220 (1989); Clothier, R.H., Quaife, Y., Ali,
I., Ruben, L.N. and Balls, M. Herpetopathol. 1:83-88 (1989).
8. Recently, we have found that mouse IL-10 will reduce B cell anti-TNP antibody
production when they are exposed early in the immune response to TNP-LPS, but
they will increase anti-TNP antibody production, when the exposure to IL-10
is delayed.
1. Long term IgM related hapten-specific memory can be initiated in the newt
by using TD and TI immunogenic carriers, but TD carriers are required for the
second challenge if memory is to be expressed. Only primary responses of the
newt are sensitive to Cyclosporine A. Ruben, L.N. Immunol. 48:385-392 (1983).
2. Anamnestic, as well as primary responses to TNP-Ficoll in Xenopus,
require T cell activity. This T cell activity appears to support or stimulate
the differentiation of the relevant B cell subpopulation from a precursor pool
that is responsive to TNP-LPS. Ruben, L.N., Clothier, R.H. and Balls, M. Thymus
8: 341-348 (1986).
1. Hapten-specific tolerance can be stimulated in Xenopus, but not
in the newt, Notophthalmus viridescens, by injection of TNBS. This tolerance
is restricted to responses that involve TNP conjugated to TI-type 2 carriers
in Xenopus. Mette, S.A. and Ruben, L.N. Cell Immunol.53:298-306 (1980).
2. Hapten-specific tolerance in Xenopus is maintained by the stimulation
of hapten-specific thymic inducer suppressor cells. Signals provided in vivo
by human rIL-2 and Con A can switch hapten-specific tolerance from unresponsivesness
to responsiveness in Xenopus. This responsivity is thymus-dependent when
TNP-Ficoll, but not, when TNP-PVP is used as the test immunogen. TNP-PVP responses,
while activating the same B cells as TNP-Ficoll, are not thymus requiring in
Xenopus. Since TNBS will not effect long-term memory responses to TNP-Ficoll,
it would appear that the TNP-specific suppressor function this reagant stimulates
is acting on the differentiation
of the relevant B cell sub-population which responds to TNP-Ficoll/TNP-PVP.
Ruben. L.N., Clothier, R.H., Merchandani, M., Wood, P. and Balls, M. Immunol.
61:235-241 (1987).
3. When a single injection of TNBS is made into premetamorphic, metamorphosing
and thiuorea-blocked metamorphosing larvae, no hapten-specific tolerance was
initiated. This absence of tolerance correlated with a lack of splenic suppressor
function in all three classes of larvae, suggesting that larvae may not have
functional in vivo suppressor function and therefore are unable to use this
mechanism in establishing self-tolerance for the larval form. This shift from
an absence of suppressor regulation of the establishment of tolerance in larval
forms to its function in the maintenance of tolerance in the adult, is in agreement
with data noted above with single haplotye disparate skin grafts made during
metamorphosis, but assayed at least 100 days later. While the establishment
of tolerance to the grafts was sensitive to IL-2 injection, its maintenance
was insensitive to the reagent. Moreover, it was dependent on suppressor function
(Horton, J.D., Horton, T.L., Varley, C.A., and Ruben, L.N. Transplant. 47:883-887
(1989).
4. When TNBS is consistently present (Stage 48 to young adulthood) in the animal's
acqueous environment, the tolerance initiated is broad-based. That is, it is
effective when TNP is presented on any of the three different classes of carriers
used. Moreover, when first applied during metamorphosis, the tolerance is effective
with TNP-SRBC and TNP-LPS, but is absent when the TNP is presented on PVP. We
suggest that this is because the B1 population required to respond to TNP-PVP,
is not differentiated during metamorphosis and therefore is unable to respond
to the exposure to TNBS during this period. Interestingly, the capacity to generate
tolerance parallels the appearrance of suppressor function in the only secondary
lymphoid organ in Xenopus, the spleen, which also develops in the last
stages of metamorphosis. No capacity for tolerance, as a consequence of exposure
to TNBS, is possible during premetamorphic life, when no splenic suppressor
function exists. On the other hand, thymic suppression may function in T cell
negative selection, since the thymus during these stages, and only in these
stages, possesses both inducer and effector suppressor functions.
5. A theory of cancer resisitance in amphibians was developed that suggested
that their resisitance may be related to their diminished capacity for altered-self
tolerance (RUBEN, L.N., CLOTHIER, R.H. and BALLS, M. Is the evolutionary increase
in epitope-specific tolerance within the vertebrates related to cancer susceptibility
? Herpetopathol.2:99-104 (1995).
1. We find that the in vivo thymic apoptotic levels are very low during
metamorphosis and therefore are not likely to be responsible for the depressed
immune activity seen at this time. Moreover, in vivo apoptosis doesn’t
appear to be regulated by the internal glucocorticoid levels in the plasma (Ruben,
L.N., Ahamadi, P., Buchholz, D., Johnson, R.O., Clothier, R.H. and Shiigi, S.
Apoptosis in the thymus of developing Xenopus laevis. Dev. Comp.
Immunol. 18:343-352 (1994); Grant, P., Clothier, R.H., Johnson, R.O. and Ruben,
L.N. Lumphocyte apoptosis in Xenopus laevis, the South African
clawed toad, during metamorphosis. (Dev. Comp. Immunol. In Press).
2. Few, if any, new antigens are being expressed during premetamorphic development.
As the animal grows, however, only modest measures may be needed to diminish
anti-self reactivity to preserve the integrity of self. A relationship between
apoptosis and the expression of new antigens is supported by our findings that
during metamorphosis in vitro thymic apoptotic rates are extremely high in comparison
and that if one freezes the animals in metamorphosis by blocking their development
using thiourea to reduce the production of thyroxine, the hormone responsible
for driving metamorphosis and the multitude of new differentiations that result,
the thymic apoptotic rate drops from 55-80% to near zero (∞7-9%).(Ruben,
L.N., Ahamadi, P., Buchholz, D., Johnson, R.O., Clothier, R.H. and Shiigi, S.
Apoptosis in the thymus of developing Xenopus laevis. Dev. Comp.
Immunol. 18:343-352 (1994). Thus, apoptosis responsible for clonal deletion
in the thymus, is driven by antigenicity. Since the peaks in apoptotic rates
do not correspond to the glucocorticoid levels in the animals, it would appear
that while corticosterone is responsible for regulating T cell functions (Highet,
A. and Ruben, L.N. Immunopharm. 13:149-155 (1987), it does affect apoptosis
in the thymuses as it does in mammals. Peripheral tolerance to TNP, in developing
Xenopus, is suppressor T cell dependent, as shown by prior treatment
with cyclophosphamide (Ruben, L.N., Goodman, A.R., Johnson, R.O., Kaleeba, J.A.R.
and Clothier, R.H. Devel. Comp. Immunol.19:405-415 (1995).
3. Adult thymocytes of Xenopus , like those of mammals, will have enhanced
apoptosis following in vitro exposure to the glucocorticoid, dexamethasone,
PHA (phytahemagglutinin), as a lectin, and TNBS (trinitrobenzene sulfonic acid)
as an antigen (Ruben, L.N., Buchholz, D., Ahmadi, P.,Johnson, R.O., Clothier,
R.H. and Shiigi, S. Apoptosis in the thymus of adult Xenopus laevis.
Dev. Comp. Immunol.18: 231-238 (1994).
4. In situ thymic and splenic apoptosis has been described over a 24
hour experimental period in adult Xenopus after injection of a T cell
(PHA) and B (LPS) cell specific lectins. The medulla of the thymus and the red
pulp area of the spleen are primarily involved in the apoptosis which peaks
around 12 hours after injection (Grant, P., Clothier, R.H., Johnson, R.O., Schott,
S., and Ruben, L.N. The kinetics and distribution of T and B cell mitogen-stimulated
apoptosis in vivo. Immunol. Letters 47:227-231 (1995).
5. A Fas-like molecule which can initiate apoptosis is present on the surface
of a variety of developing and adult tissues, including the thymus and spleen,
of Xenopus (Mangurian, C., Johnson, R.O., MacMahan, R., Shiigi, S., Clothier,
R.H. and Ruben, L.N. Expression of a Fas-like apoptotic molecule on larval and
adult cells of Xenopus laevis. (In press,Immunol. Letts.).
6. Adrenoceptor ligation will strongly enhance or reduce concurrent apoptosis
stimulated by either the calcium ionophore, A23187 or the phorbol diester, PMA,
in accordance with the class of receptor stimulated and the length of time of
exposure to either the alpha-2, clonodine, or beta, isoproterenol, agonist used.
Apoptosis induced by dexamethasone was resistant to modulation by both adrenergics
(Haberfeld, M., Johnson, R.O., Ruben, L.N., Clothier, R.H. and Shiigi, S. Adrenoceptor
modulation of apoptosis in splenocytes of Xenopus laevis in vitro.
(In press, NeuroImmunoModulation).
1. Helper function can be demonstrated during larval Xenopus development
but, it is aberrant during metamorphosis. Ruben, L.N., Welch, J.M. and Jones,
R.E. In " Development and Differentiation of Lymphocytes of Vertebrate
Lymphocytes ". Ed. Horton, J.D., pp.227-240, No. Holland Biomed. Press,
Amsterdam (1980).
2. An internal MLR, leading to the release of thymus replacing factor during
metamorphosis, may establish a "bypass" of the usual requirement for
helper function in Xenopus. Jones, S.E. and Ruben, L.N. Immunol. 43:741-745
(1981).
3. Inducer and effector suppressor cells are functonally inhibited and become
anatomically compartmentalized during metamorphosis of Xenopus. The thymus
of pre-metamorphic larvae, unlike that of adult toads, contains both inducer
and effector suppressor functions. Kamali, D., Ruben, L.N. and Gregg, M. Cell
Diff.18:225-231 (1986).
4. Responsivity to TNP-Ficoll varies with developmental stages and internal
endocrine environmental changes with age. Ruben, L.N., Clothier, R.H., James,
H.S. and Balls, M. Cell Diff. 14:1-5 (1984).
5. Corticosteroid will inhibit a TD response in an adult lower vertebrate. Ruben,
L.N. and Vaughan, M. J. Exp. Zool. 190:229-235 (1974).
6. The number of corticosteroid receptors/cell of splenic lymphocytes during
metamorphosis, when serum titers are high, is 10x the level found in young adults.
Pre-metamorphic larvae will not bind the hormones.Since corticosteroids will
inhibit T cell mitogenesis, helper and suppressor functions, it seems likely
that T cell functional deficiencies during this period are mediated by corticosteroids.
Marx, M., Ruben, L.N., Nobis, C. and Duffy, D., In " Developmental and
Comparative Immunology ", Ed. Cooper, E.L., pp 129-140. Alan R. Liss Inc.
Publ., New York (1987).
7. Corticosteroid is responsible for the inhibition of inducer suppressor function
during Xenopus metamorphosis. This inhibition can be relieved by injection
of metyrapone, human IL-2, Con A or human IL-1in vivo. Thus, this T cell immune
deficiency seems to be due to inhibition of Mø secretion of IL-1 and,
in turn, too little IL-2, by high endogenous corticosteroid titers
Highet, A. and Ruben, L.N. Immunopharm. 13: 149-155 (1987).
8. Alpha and ß adrenergics effect the capacity of immunized adult Xenopus,
but not of newt, B cells to bind antigen differentially. Hodgson, R.M., Clothier,
R.H., Ruben, L.N. and Balls, M. Eur. J. Immunol. 8:348-351 (1978).
9. Human IL-2 will substitute for helper function in Xenopus. Its capacity
to do this will last for three hours prior to antigen challenge. Ruben, L.N.
Immunol. Letts. 13:227-230 (1986).
10. Human IL-2 and Con A can stimulate responses to sub-immunogenic dosages
of TNP-Ficoll. Ruben, L.N., Barr, K., Clothier, R.H., Nobis, C. and Balls, M.
Dev. Comp. Immunol. 9:811-818 (1985).
11. Murine and human IL-2 can substitute for the thymus requirement in TNP-Ficoll
responses in adult Xenopus. Ruben, L.N., Clothier, R.H. and Balls, M.
Cell Immunol. 93:229-233 (1985).
12. Signals provided in vivo by human IL-2 and Con A can switch hapten-specific
tolerance from unresponsivesness to responsiveness in Xenopus. This release
from tolerance is thymus-dependent when TNP-Ficoll, but not when TNP-PVP is
used as the test immunogen. TNP-PVP responses, while activating the same B cells
as TNP-Ficoll, are not thymus requiring in Xenopus. Ruben. L.N., Clothier,
R.H., Mirchandani, M., Wood, P. and Balls, M. Immunol. 61: 235-241 (1987).
13. Xenopus splenocytes display constitutive molecules which bind mouse-anti-human
IL-2 receptor antibody specifically. Il-2 will compete with this binding. Moreover,
the number of cells able to bind both the anti-receptor antibody and IL-2, as
well as the number of receptors/cells can be increased by lectin activiation
of the freshly biopsied spleen cells in vitro. Langeberg, L., Ruben, L.N., Malley,
A., Shiigi, S. and Beadling, C. Immunol. Letts. 14: 103-110 (1987).
14. The Xenopus spleen cells which are PHA activatable with regard to
IL-2 R receptor density and which bind rIL-2 are related to helper T cell function
in cytotoxic and humoral immune responses, since it is removed by N-CH3-N-Nitrosourea
(NMU) injection. NMU is selectively lymphotoxic in the toad and specifically
removes these two functions. Panning spleen cells with a mAb to Xenopus
IgM showed that those cells which bear constitutive IL-2 receptors and bind
rIL-2 are equally abundant in the T and B cells populations of freshly biopsied
Xenopus spleens. Langeberg, L., Ruben, L.N., Clothier, R.H., and Shiigi,
S. Immunol. Letters 16: 43-48 (1987).
15. Human IL-1, Con A on sepharose or agarose, but not soluble Con A or soluble
or particulate WGA or human IL-2, will substitute for carrier primed helper
function in the newt. While anti-human IL-2 receptor antibody will bind specifically
to freshly biospsied spleen cells, Il-2 will not compete with this binding.
The shared lectin specificity suggests that T-like cells are responsible for
helper function in this animal with an " immature " thymus. The lymphokine
and Con A data suggest that while human IL-2 is unable to affect the cells of
this species, Con A and human IL-1 can initiate the stimulation, perhaps through
the stimulation of autologous IL-2 production. Ruben, L.N., Beadling, C., Langeberg,
L, Shiigi, S.and Selden, N. Thymus 11: 77-87 (1988).
16. The monoclonal mouse-anti-human IL-2 recedptor antibody that we have used
recognizes molecules of the cell surface of Xenopus lymphocytes that
specifically bind rIL-2 and are essentially the size as the Tac molecule (p55)
Ruben, L.N., Langeberg, L., malley, A., Clothier, R.H., Beadling, C., Lee, R.O.
and Shiigi, S. Immunol. Letters 24:117-126 (1990).
17. During metamorphosis, when T cell functions are impaired by a high glucocorticoid
titer, T lymphocytes also have reduced capacities for the expression of IL-2
receptors, upon exposure to PHA, and to generate autologous TCGF. Thus, we have
suggested that during this developmental period, when adult cells and antigens
are being expressed within the larval body, T cell functions are impaired by
glucocortioid-imposed anergy and that this is what is responsible for the prevention
of immune self-destruction (Ruben, L.N., Scheinman, M. A., Johnson, R.O., Shiigi,
S., Clothier, R.H. and Balls, M. Mech. of Develop. 37:167-172 (1992).
18. Mortality in developing larvae, particularly during the metamorphic period,
can be manipulated by using IL-2 along with an antigen. Thus, it may be possible
to lower the amount of IL-2 used in experimental cancer therapy by adding an
antigen in conjunction with it (Ruben, L.N., De Leon R.T., Johnson, R.O. and
Clothier, R.H. Interleukin-2-induced mortality during metamorphosis of Xenopus
laevis. Immunol. Letts 51:157-161 (1996).
19. In vivo thymic apoptotic levels are very low during metamorphosis and therefore
are not likely to be responsible for the depressed immune activity seen at this
time. Moreover, in vivo apoptosis doesn’t appear to be regulated by the
internal glucocorticoid levels in the plasma (Ruben, L.N., Ahamadi, P., Buchholz,
D., Johnson, R.O., Clothier, R.H. and Shiigi, S. Apoptosis in the thymus of
developing Xenopus laevis. Dev. Comp. Immunol. 18:343-352 (1994);
Grant, P., Clothier, R.H., Johnson, R.O. and Ruben, L.N. Lumphocyte apoptosis
in Xenopus laevis, the South African clawed toad, during metamorphosis.
(Dev. Comp. Immunol. In Press).
20. Few, if any, new antigens are being expressed during premetamorphic development.
As the animal grows, however, only modest measures may be needed to diminish
anti-self reactivity to preserve the integrity of self. A relationship between
apoptosis and the expression of new antigens is supported by our findings that
during metamorphosis in vitro thymic apoptotic rates are extremely high in comparison
and that if one freezes the animals in metamorphosis by blocking their development
using thiourea to reduce the production of thyroxine, the hormone responsible
for driving metamorphosis and the multitude of new differentiations that result,
the thymic apoptotic rate drops from 55-80% to near zero (∞7-9%).(Ruben,
L.N., Ahamadi, P., Buchholz, D., Johnson, R.O., Clothier, R.H. and Shiigi, S.
Apoptosis in the thymus of developing Xenopus laevis. Dev. Comp.
Immunol. 18:343-352 (1994). Thus, apoptosis responsible for clonal deletion
in the thymus, is driven by antigenicity. Since the peaks in apoptotic rates
do not correspond to the glucocorticoid levels in the animals, it would appear
that while corticosterone is responsible for regulating T cell functions (Highet,
A. and Ruben, L.N. Immunopharm. 13:149-155 (1987), it does affect apoptosis
in the thymuses as it does in mammals. Peripheral tolerance to TNP, in developing
Xenopus, is suppressor T cell dependent, as shown by prior treatment
with cyclophosphamide (Ruben, L.N., Goodman, A.R., Johnson, R.O., Kaleeba, J.A.R.
and Clothier, R.H. Devel. Comp. Immunol.19:405-415 (1995).
Maintained by the Reed College Biology Department
Last Modified 8/20/02
Questions/Comments to bioweb@reed.edu